MCAT Rates & Enzyme Kinetics (Part 2)

[MUSIC] Welcome to the Jack Weston MCAT podcast with your hosts, Michael Malayka and Molly Huda. [MUSIC] Hello everyone and welcome back to the Jack Weston MCAT podcast with your host, Mike and Molly. This is a pretty special episode because it's actually a part two of a previous episode. We started off last episode talking about kinetics and enzymes and there's simply too much to talk about in that realm that we said pause. Let's do round two, right? Right. So what are we talking about with enzymes and kinetics today, Mike? Right. So we already covered kind of like the basics of rates, right? That's like part one was all about what is right and how it's so different from thermodynamics. Thermodynamics. Different, totally different planet. And if you're like, why are they talking about planets right now, go watch that episode. Go watch, baby. I guess yeah, if you, this is, I think one of those only episodes that we've done so far where if you actually haven't watched the first episode, you should actually pause and watch the first episode. So like the other ones, like you can kind of jump in about too much contacts, but this is one where having watched episode one, go watch episode one of this, the one that's just previous. Or if you're already very comfortable with rates and what an enzyme is, then it's fine. If you've reviewed this material before, but if you're coming to us to learn content, maybe not for the first time, but maybe for the first time in a long time, go watch it. It will be worth their time. Definitely worth your time because we're going to really be building off because yeah, we've covered right we're not going to cover too, too much in depth today. But again, it's all about speed, it's all kinetics, not about thermodynamics. We've also covered some things that can impact right the different variables that can impact rate. We talked about the arenas equation that kind of shows us. We've talked about our rate equation that kind of shows us how reacting concentration different orders of reaction. If I'm saying words and you're like, wait a minute, I don't know those words, go watch the last episode. First order second order zero, the order. How does reacting constant substrate concentration impact it? But then we also talked about the enzyme, like how enzymes actually impact the rate because we know the impact rate not favorability. We made that distinction very, very clear, but it's not favorable to start adding an enzyme, I'm not going to do anything. All it does is speed things up and how it does that? Lowering the activation energy making it so you don't have to invest as much energy in to get over that hump to get over that kind of that hill of that high energy intermediate. And then we talked about what actually is this enzyme, the big ball of amino acids, right, the big protein, the different sites that are important. And so we're really jumping into on enzymes and actually how we study how enzymes impact rate. So Molly, we're going to be chatting about mechilles mentium. We're going to be telling about inhibitors. I'm excited. And line we were birds. And line you. I know you really like the line of irrequence. And again, something that so many people really dislike. So I'm happy that we're going to talk about it and someone who's as passionate about it as you are is going to be chatting about it. Well, here's the thing. I feel like I feel like there's this misconception with a lot of at least students that we see in classes. I assume the supplies podcast listeners too, but when students come to class, I think there's this expectation that the people that are teaching the classes or hosting the podcast have just. Oh, they were like born with this knowledge, like they never had to struggle for this. But that is so not accurate, right? And all the things that I personally get really pumped to talk about are the things that bottled my mind when I first started, right? Or the things that I had trouble making sense of I had trouble memorizing again, not a memorization person. So the things that bothered me initially are the things that now I love. Right, but I feel like it's valuable to point out that no one was born with all of this knowledge. And I mean, you don't even have to be this walking encyclopedia to get a great score. Anyways, maybe you weren't born with it, but you also don't need to be that, you know, absolute chemist, physicist, genius in order to score well. Right. That's the beauty of the MCAT. So yes, we will talk about Limey Rubirk. In fact, I love all of enzyme kinetics. I'm super excited to get into this. But I feel like as you mentioned, we covered a lot last episode. And there's one thing in specific that I feel like is super relevant to what we'll talk about today, especially when we talk about inhibitors later. And that's just the sites on an enzyme. Right. So to set our stage and enzyme is a protein, big long chain of amino acids. This is a protein that's specialized to catalyze a reaction. Like you said, it's not a thermodynamic thing. It's not a thermodynamic beast. I think so. I called it last episode. It lives in kinetics land. Right. It's very much based on speeding things up. It does not have any effect on equilibrium. Like you mentioned. And non spontaneous reaction, it can't suddenly cough up the energy to get that to happen itself, at least. What it can do is reach out and grab onto a substrate. Right. That is going to enter the binding site. That's part of the active site. And then we will see some sort of catalytic reaction. And then the time is helping speed up the reaction, not by being an active participant, but by helping the uncomfortable transition state where your half enzyme and half substrate and half product become more stable. Okay. So when we talk about an enzyme, the active site is really where the magic happens. That's where the substrate is going to bind and undergo that catalytic reaction. And then there are other sites all over that gigantic often globular protein is big ball twisted ball. Sometimes multiple peptides in there too. Any other site on that enzyme that's not the active site. It's not where the substrate binds could theoretically act as what we call an alistere site. Right. So all of the other steric is place. So allosteric is another place on the enzyme that's not binding the substrate. This sounds abstract now, but it's going to come into play very specifically with inhibitors. So that's the one part of last episode that I want to make sure even if you listen to that we're comfortable with that idea in active sites versus an alistere site. All of inhibition is built on in a way. But right, the rest of it go watch the episode. Today, I think the best place to start to pick up where we left off in kind of joining these two topics of enzymes and kinetics is with McAleous mentan. I love McAleous mentan. I love McAleous. And I'm excited because I feel like we have very opposite things that excite us. And I know that you really love the math of McAleous mentan and the research of McAleous mentan. Yes. Yes. So that's the story of course of course again, because I resonate so much with like these are things I didn't understand. So I it never really made sense at first. I was so confused. I think the biggest misconception about McAleous mentan plus is all just one experiment. Right. It was all just happened just one time. But no, let's take us back to actually what happened. So way, way back, I don't have a date. But we want to study how substrate concentration impacted the reaction rate. We want to see if I change substrate concentration, how would I impact the reaction rate of an enzyme catalyzed reaction. We understood zero or third or first or second order. But we want to see, okay, how does an enzyme change things. So what they did. So McAleous mentan actually don't know if these were two people one person. This I don't know. Well, I think it was two people, right. But what they did was that they took a small file, the small file and added a measured amount of enzyme, measured amount of enzyme to that small mile. Let's say 10 molar. And then if there's 10 molar enzyme in this small vinyl in this solution and there's no substrate, would there be any reaction. No, there's nothing to react. But what they did was they added a very small amount of subject, very small amount. And then they measured the initial reaction rate and then stopped the experiment. Right. Only the initial reaction rate imploded it. They then did this entire thing. They threw out that file. They were like, okay, we're done with that. Made a whole new file had 10 the exact same concentration of enzyme and then added a little bit more substrate, measure the initial reaction rate and then stop the experiment, plotted that and they did that over and over and over and over again. What they found was that at first, at first when they started adding more substrate concentration, the initial reaction rate would get go higher. It would lead to having much faster initial reaction. Of course, because now these enzymes have all this substrate around that they can grab on that that they weren't working now they can. But it's very strange thing happened where as they started adding more and more substrate concentration, the increase in initial reaction rate will get. It would still go up, but won't go up by as much and eventually they got to a point where they started adding substrate concentration and that wouldn't impact their reaction at all. The reaction would be the exact same, even though they doubled tripled quadrupled the amount of substrate concentration. And then they plotted this data and this is where we get the mechalism and graph this hyperbolic curve. I want to say that word because that's a word that we do need to know. It's this hyperbolic shape where at first it goes up as we increase substrate concentration that's our x axis. That's what we're increasing. We're increasing it and our y axis is the initial reaction and we see that at first is going up and up but then it just levels out no matter how high you go. Now this is the idea of Michealis Minton and why it's in my opinion in my thoughts is very important to know how we got the curve because if you think this is one experiment it makes no sense whatsoever. No sense what how are you changing stuff right how are you having changing initial reaction rates if it's just one experiment no this was multiple experience that again and again and again and so I think it's so cool because now it makes sense why it looks the way that it does and from this graph there's some important points but kind of before we talk about the really important parts of this graph I like kind of hitting on the y-axis because there's something I said very intentionally a lot which was the initial reaction right right this wasn't just any reaction right didn't wait five seconds ten seconds or waited even to equilibrium they only measured the initial reactionary of these experiments but then the fun used to be why why did they do that it's because when they when they want to see how substrate concentration affected that forward reaction but if you wait five ten fifteen seconds what happens is you form product and the reverse reaction occurs so if you wait five ten seconds you are only seeing the net reaction when you're looking at these vials so you're not actually measuring the forward the forward reaction anymore you'd be measuring the net of the forward and the reverse so there may be the forward reaction is ten molar per second but if you waited five ten seconds the reverse reaction might be two molar per second so you'd measures actually eight which would be incorrect when reality was ten and so this is that really important note that this is not just any reactionary on the y-axis it's the initial reactionary it's important because of that reverse reaction again I nerd about this I get I'm starting to get rambly because I this love talking about this but this is kind of when we look at a mccalesment and it's important to know how they got that graph in the first place yes and I love the history lesson I tend to go way way back in evolutionary biology and I like that you get excited about this and I like that you point out the initial reaction rate and also just to really re-emphasize this there's always the same amount of enzyme in it we're not changing the amount of enzyme that's in that vial it's always at a set concentration and then we're changing substrate concentration right and we've got on the x-axis of this graph right it is substrate concentration so whatever substrate we're interested in if it's a multi-substrate reaction right we look with respect to each one individually so you have substrate concentration on the x and then reaction initial reaction rate as you said on the y so cool to think of how wonderfully frustrating that must have been to plot but again we end up with this curve and for me I love connecting this to rate law because it's it's really really cool to think of what happens we start out with basically a linear relationship as substrate concentration um increases when we consider the shape of the curve right it's it's almost linear right increase substrate and reaction rate goes up a lot and then as you said it starts to kind of level off and actually now you might add more and more substrate maybe you had twice as much substrate but you only increase a little bit okay you don't increase double um like you would earlier right it levels off and then it gets flat it's really cool back when we were talking about first second and zero-th order reactions right we start off linear ish right that's a first order reaction right as we continue it levels off to a zero-th order reaction right because changing substrate concentration does nothing to rate and a lot of students ask why why does it level off what is the meaning of that the slope of the graph changing and I think the important key piece here is you're not adding any more enzyme right I personally the thing that makes me uh well one of several actually that makes me so excited about enzyme kinetics is we think of this maximum rate that we can get the fastest that that rate can get as this one objective number that never changes like ah the vmax of this reaction is however many reactions per second like we have it in our mind that you can't never change that um and vmax again would be the point that we don't really cross when we increase substrate concentration and we keep increasing it we we level off right we get horizontal parallel to the x-axis we never go higher in our initial reaction rate um and that actually happens when our enzyme is completely busy and I love this because I think of I think of enzymes as you know and Mike's gonna laugh because I I always humanize my biochemistry but like I think of an enzyme as a as an individual person right and enzyme is capable of carrying out a task right you can if if you're doing nothing and you get a homework assignment you could do it right no problem right if you suddenly got two homework assignments okay you could still probably do it right three okay this is more now but like you could still do it there's a point that you you cannot work that fast like there is a inherent limit to how much you can do and how fast you could do it every human being has limits we only have 24 hours in the day there is a limit um and enzymes are the same way as an individual they have their own maximum speed of work this is as fast as I can physically work that is not v max right for an individual enzyme this is actually what we call K cat or the turnover number it's the maximum rate of reaction for one enzyme and that is that is the that's K cat is unique to the enzyme period one enzyme can only work so fast when we look at v max though that can actually change we can change v max and we can imagine this if right considering that test tube reaction if we only had one enzyme in there right you can map that whole reaction but you're going to reach max the absolute maximum pretty quick because you only have one enzyme and it can only do so much what would happen if suddenly I had two enzymes in that test tube or four or six or eight you now have in the same period of time two or four or six or eight times greater capacity to catalyze that reaction because every individual enzyme can work that fast right and that is the basis of v max v max is objectively the rate how fast one enzyme can work times essentially how many enzymes you have enzyme concentration but we can change v max based on how many enzymes were there so hypothetically in your experiment as we were describing it right we we reached this point where to our little test tube with the same amount of enzyme that we've always had you can add substrate at double twice or double triple quadruple the amount that you had in the previous experiment and it does nothing to your v max if instead you doubled the enzyme concentration you would be able to achieve that fast growth again you can you can move your v max right you could change the shape of your graph because at v max you have individual enzymes that are drowning in substrate right imagine you're a student and you've got 600 home work assignments you can't snap your fingers and do all of them you can work as fast as you can you can keep chipping away at things but at the end of the day there are limitations right the only way to change v max is to change the amount of sub or amount of enzyme that you have so that there's more hands that can complete in that case the assignments but really this you know catalyze the substrate reaction at a greater extent so I love that stuff I'm the way you explain like how you preposing with k capric I think it's really cool because yeah you're totally right people we think that again I know I thought about like first like this like v max is just this salty thing that can't move because that's what we're used to in a mccalesment and graph we have to realize yeah in mccales meant in graph enzyme concentration would stay the same throughout but if I change the amount of enzymes it's like I have more workers I've got more people doing work v max is just the total work that's happening between all of our enzymes right but yeah if you just make more workers yeah more total work is going to happen um and then thinking about k cat is like the maximum of a single enzyme I think if you define k cat in that way well just of course makes so much sense and again this is I think going back to this idea of not memorizing because I'm sure a lot of people here probably we're listening probably memorize the k cat equation k k as v max over enzyme concentration which I hate also right which so but I think if you take it take it take it as weight k cat wait v max is just k cat times the amount of enzymes enzyme concentration right like you don't you don't have to memorize the other way yeah exactly the equation just your range a little bit but it just more is mid just long make a lot more sense right it just it's more intuitive in that way um and that's what I always kind of like I'm actually a really big fan of rearranging equations to make them intuitive to help me understand them rather than memorize them again it might not be the way that my equation sheet that I memorized kind of wrote them out but if you can make like rearranged an equation to make it intuitive rather than memorizing do it as long as you're not changing the equation yes as long as the math is the same the math don't like start adding variables like no plays don't do that but if it's the same equation yeah rearrange it I do that for the I mean we haven't talked about the index of refraction equation like the n equals see over yeah I do that one I rearrange that one to make it make more sense to me um but yeah we can talk about that another day I don't know the time yeah but that's absolutely this v max right and kind of going back to the main Achillesmending curve was that maximum initial reaction rate. When McAleous and Menton kept on doing the reaction, adding more and more subs, say, that was that initial reaction that did not move. No matter how many, how much more substrate they added, those enzymes couldn't work any faster. Right? Yeah. And so that, the max is a very important value when it comes to McAleousmending. But there's another important value we do need to talk about, which is KM. Yeah. And so I want to preface when it comes to Ks, right? Again, I did not make the rules of chemistry. There was somebody who really loved the letter K in chemistry. They love that letter because we've talked about a few case already. Like over the last, you know, however many podcasts episode, we first talked about big capital K K equilibrium, concentration of products, concentration of reactants. We've just talked about the rate K, the slower case K, which is all about the rate constant. Now we can talk about another K, which is KM, the McAleous constant. And now here's a kind of a big new one thing. It is a capital K. It is capital K, but it is not an equilibrium constant. Yes. It's the only exception to that rule. Big K small subscript. It's a type of equilibrium except KM. It's the only one. And this is the one where I notice a lot of students when they're like doing their passages, they see capital K. The first thought is ill. It's like KM. That should not be the first thought. It's like that saying, you know, when you hear hoes, think horses, not zebra's. You see a big K, you should first assume it's probably an equilibrium constant. Unless there's something to tell you that this is a McAleous constant. Don't first jump that to McAleous constant because that's the exception, not the norm. But let's talk about what this KM value is. So KM is defined as the substrate concentration to reach one half V max. Now I want to climb through them. KM is not one half V max. It is not one half V max. Because that's like you're a lot of people. It is not a reaction rate. KM is the substrate concentration that you need in order to reach one half V max. Now this idea is very important because we have to realize that it based on that definition, what would be the units of KM? It would be substrate concentration, molar, something like that. It'd be molar, some millimolar and animal or whatever. It's a concentration. It's not a rate. It's not inverse molar. A lot of equilibrium constants are inverse molar. But that's just based on the equation. But no, if you see a capital K with the constant with a units of molar, now this makes you think that this might be a KM. But otherwise, like if it has no units or units that are non one over or that are not molar, it's an equilibrium constant like flat out. So KM defined as the substrate concentration to reach one half V max. But then the question needs to become, why do we care about that? That's the point. That's the point. Like, okay, you've told me how much substrate you need to get halfway there. Like, okay, why is that important? Because when you think of KM, the word that you come to mind is affinity. That's the word. And what do we mean by affinity? Affinity is just how we kind of frame how badly does this enzyme want to bind the substrate? And how much does it like binding to the substrate? And we use KM to tell us that information. Because imagine, let's imagine with me, Molly, I need just like, for imagining right now, imagine I just need to add a little bit of substrate. Like I put a drop of substrate in and then my reaction rate shoots up to one half V max, right? Shoot. So did my enzymes, when they saw, were they like pickier? Did they grab it right away? Oh, they grabbed it. They were very excited about it. They were, I added just a little bit of substrate and they shot up. But I want you to contrast that to if I had had so much substrate in order to get to that one half V max where my enzymes excited to bind to that substrate or were they like kind of picky about it? They're pretty picky. They were really picky. And this is how I like to frame KM in terms of affinity where a low KM means a low amount of substrate to reach one half V max. Tell us that is a high affinity. Low KM is high affinity. They have this inverse relationship, whereas a high KM, a high amount of substrate to reach one half V max is a low affinity. They're gonna pick you. They don't don't want to bind to that substrate as much. You know, you know, you know, I always got to humanize things. Of course. I always explain this as a procrastinator. There are two kinds of students. There are the students during syllabus week that first week they get one assignment that's not due for two months and it's done the next day. Like they have very, like everyone knows at least one of these people. It's not really me either. But like the they are they're excited to do the work. They're excited. This is a class they care about. Like I don't care that it's not due for a very long time. I'm gonna do it now. Right. I have a very low amount of work to do. And my deadline is not very present, but I'm very drawn to the work and I'm gonna do it. Right. That is a very self motivated, right. High affinity enzyme. That would have a very low volume of work necessary to get it to one half of its max speed. Right. So again, low KM means that you only need a little bit of work or of substrate in order to get you moving fast. Right. On the other end, right. We're probably several of us fall. Including myself. Accident leaders. Right. You get through syllabus week. You have an assignment that's not due for a while. You're like, okay, I don't care. Then you get another assignment the next day. That's doing like two weeks. You're like, okay, still not really going to do anything. And you get another assignment. Right. And it kind of piles up. And then you look and you're like, okay, actually, there's a lot to do. Now I'll start working. Right. You do not have a very high affinity towards that work. Again, the KM in that case, your affinity is going to be or your KM, I should say, your KM is very high because you have to have a lot of work or a lot of substrate around you to get you moving. Right. And keep in mind the reason why we say one half V max is because every enzyme and again, different concentrations to KM can change. Right. So rather than saying like how much substrate is necessary to get you at X amount of reactions per second. Right. That's some, some enzymes may never move that fast or some enzymes may move that fast, even like almost at rest. We use one half of that V max of that reaction so that we have a way to describe affinity that's not really dependent on whatever that V max value is. Right. It's basically. If V max changes that one half V max is going to change. Exactly. Yeah. So it's allowing us to describe the affinity. Again, if you have a very low KM, you need very little substrate around to get going and to start working. You can get to one half V max. That's a very high affinity and the reverse history of our procrastinators. High KM, very low affinity. And that kind of draws us nicely to the idea of catalytic efficiency. Oh my goodness. I get so heated about this. Oh, I'm excited. Be calm. No, honestly, no, you go. I get no, you go, please, please. Let me just lay the groundwork. Okay. Because I want to see you, you know, crash out of the crash. I'm about to crash out. I'm about to. Okay. Catalytic efficiency is this equation that we can use to help us understand how well an enzyme works in essence, right? KM and V max are going to be specific values, a substrate concentration or a reaction rate. And those are hard to compare between reactions, right? In between different enzymes that do different things. So we have this equation, like efficiency that is going to take into account KM for that enzyme and also K cat, not V max, which is and concentration dependent, but that K cat, that's how fast can this one enzyme move, right? Or else you'd see different efficiencies for, you know, solutions that had different concentrations of enzyme. No, no, K cat and KM, right? I always assume in order to help you remember, it's a ratio. If I want a very efficient enzyme, I want a super high K cat, because that means you move very efficiently or very well, very fast. I know I know very fast. You're able to catalyze reaction fast. And then a low KM, because you want it to be high affinity. So catalytic efficiency, if you want to max it out, K, K cat is on top, KM is on bottom. That's the basics. Now let's hear the crash out. I'm about to crash out because I want everyone listening to never, I'm about to make a statement and you are never allowed to make this mistake. Anyway, listening K cat is not equal to catalytic efficiency. K cat is not the catalytic efficiency. I have, oh my goodness, we've been doing this for years. I hear that all the time, right? Oh, right. Oh, K cats, the efficiency, absolutely not. K cat is the turnover rate, right? It's the reaction rate of a single enzyme. The efficiency is how efficient that enzyme is. But the way I like to prep is like, let's take a scenario, let's say I have an enzyme that, you know, can, once they bind to the subject, can move really fast, but takes forever to them for them to bind. Is that a very efficient enzyme? No, let's say it on the flip side. Let's say I've gone and done that. Bines really, really well, but takes forever for them to actually catalyze that reaction. Is that efficient enzyme? No. So you have to take both the speed of turnover, that turnover rate, and the affinity into account. And that's where catalytic efficiency comes from where you have to take both the K cat and the KM into account. But again, I always want to write teacher class and make everyone in the chat say this K cat is not the catalytic efficiency. Don't, please don't make that mistake. I'm about to crash out if I see that mistake one more time because I just see it way too way too often. Yeah. Yeah. It's also funny as an, or not funny. It's, it's wild as an instructor because over the years, we've worked with so many different students. and we see students come in that pretty much for the most part have very similar issues and misconceptions to begin with. So it's like we see this problem again and again and again, always with different students, but we see the same pattern play out. And yes, I've seen that many times too. You're all thinking, Mike's just being exaggerated. No, when you see the same mistake hundreds of times, you're like, I don't want to ever see this mistake again. So you've all been warned. You've all been warned. KKOT is not the Catholic efficiency. That's that's nice and ominous. You've been warned. Okay, so let's steer clear of the catalytic efficiency discussion now that we've got threats on the table. Let's talk about probably the reason why some of you guys are listening, which is Lineweaver Burke. And this is kind of it's similar. It's bringing back electrochem vibes from our previous episodes. How many people are scared of Lineweaver Burke? Just like so many people are really scared of electrochemistry. But really Lineweaver Burke is so easy once you've laid the groundwork that we already have. Somebody just got triggered right now, but just by saying that they're like easy. What do you mean? The groundwork is laid right now. And if you don't believe me, let's prove it. Right. Right. Lineweaver Burke plots have, and I'm going to hold everyone's hand while saying this, the exact same information that's in a McAleous Menton plot. Except let's say if you want to find the KM, you don't have to look at the X or the Y axis that has the Vmax, right? You don't have to find the place where it flattens out and then carry it over, find Vmax, and then calculate what's one half of Vmax go here and then follow that over, find where it intercepts the curve and then drop it down to the X axis to find the M. That's a lot of steps. And I'm like, it's a lot of steps. Yeah. It is a lot of steps. It's not that hard. Students can most students comfortable with McAleous Menton, but it's a lot of steps and there's a lot of opportunity to go wrong. Lineweaver Burke, what they have done is take the McAleous Menton equation that plots that curve, right? They've taken that equation. And on both sides, they've done one over. It's what we call a double reciprocal plot. And I know there are people that are like, why the heck would you do this? That makes things way more complicated. Instead of X axis being substrate concentration and Y axis being that initial reaction rate, it's now one over substrate concentration and one over the reaction rate or the initial reaction rate. Is that annoying? Yes. But it also gives the most convenient plot type. It's a line a straight line. It's just straight line, right? It's a straight line. And the beauty of Lineweaver Burke. Again, everything that we just talked about with McAleous Menton is exactly the same. Bmax, KM, that all the values we talk about, the ideas we talked about, it's all identical. Lineweaver Burke is the same thing, except you flipped both sides of the graph. Now we get a line and rather than having to go through all those steps to see Bmax and KM, you can actually see them. They are the X and Y intercepts. Right. It's beautiful. I love it. It simplifies it so much because you know exactly where to look. You don't have to try to do all these calculations. You just know if I need Vmax, this is where I go. If I need KM, this is where I go. And it makes life so much easier. So let's say Molly, I need to find the Vmax, right? You just said it makes it so easy to find the Vmax. Yes. Where would I find that? So you'd think about it first, right? The X and Y axis, right? X is one over substrate concentration. Y is one over the initial reaction rate, right? Vmax is that going to relate more to the reaction rate, the Y axis or the X axis, one over substrate? This is a tough one. I'm going to have to say the Y axis. It's a trick question, but yes, the Y axis. Right. And I just said that Vmax and KM are described by the intercepts. So we're probably going to be looking at the Y intercept. The only thing that we need to be really careful of is that the Y axis is actually one over the reaction rate. So what we're going to see, the Y intercept is one over our Vmax, okay? But that's it. Right? You find it on the graph. And I mean, you don't super, or you don't very frequently see them asking you to calculate Vmax from a line weaver Burke. Most often when you see these, they're going to be used for inhibitors. And to be honest with you, inhibitors and line weaver Burke, very scary at the beginning because you're just trying to memorize Oh, an uncompetitive looks like this. The shapes once you clue in to X axis is describing the KM. Again, it's one over the KM and actually based on the fact that the line crosses both quadrant one and two. It's actually negative one over the KM. But you can't have a negative substrate concentration. You drop it, right? The fact that the X intercept represents KM and the Y intercept represents Vmax, you can do those line weaver Burke questions with inhibitors super fast. And that's what they're more likely to do for you. But that's how you'd find Vmax from line weaver Burke. And the exact same process is true of KM. It's just find the X intercept one over, of course, technically it is to the left of zero on that number line in the graph. So you drop the negative sign again. No negative substrate concentration. Right. It's very cool when you look at the data planted on the leader of a Burke. It's all in the positive quadrant. Like there's no actual data points in the negative area. They've just extrapolated the line. Like that's the kind of key. They've just extend the line down. It's only like they had data points on that side of the deaf. If you actually looked at the data points, they're all on the right side. Yeah. Yeah, because you can't have negative substrate. Right. You can't. That's impossible. But that doesn't make sense. That doesn't really make sense. But yeah, but again, really realizing it again, this is where I like the math about it because all they've done is they actually took the mechalesment in equation. Now, if we can put it up on the on the YouTube video, we will again, I'll leave that to the editors, but I'll just say it verbally as well from the mechalesment in graph we talked about before. Of course, if there's ever any plot, we have to have an equation for that plot. So the mechalesment in equation is initial reaction rate, which is that y axis again on the ke-mechalesment in plot. We're not talking when we remember it right now. Mechalesment in is v not equals. It's a fraction on the top part and the numerator of the fraction. It's v max times substrate concentration and the denominator. So that's over K m plus substrate concentration. So this is our mechalesment in equation. And that's what we generated from them. Mechalesment in front of all those experiments. We drew a line where like, okay, let's what's the equation of this line? So we can actually calculate using these variables what v not is. But when they took that double reciprocal, all they did was take the inverse of that plot. So they just lit both the y and just, both sides of the equation. So it became one over v not equals K m plus substrate concentration over v max times substrate concentration. They did some great math there. And I'm gonna, I will never teach this in class. I feel like doing these are places that I'm like, I hope somebody, I hope we can get a picture of this on the video. But what they ended up finding was an equation, the linear of a bird plot equation, which is just y equals m x plus b. That's how they plotted it, which is just a little equation of a line. And so I like processing about that. Again, I feel like I'm like, I want to write things down, but I can't, but I guess it is, it is good to know what the linear of a bird plot equation is. But I mean, you can just learn that from the, from the graph, if you kind of think about it, so the y is the y axis, one over v not equals m, which is the slope. And now we haven't talked about that, but that's K m over v max. And so that's also, if you don't want to memorize, just do rise over run from left from x intercept to y intercept. You calculate it, you don't need to memorize. You don't need to memorize it. So that m is just the slope, which is just rise over on, which is K m over v max times x, which is your x axis, which is one over substrate concentration plus b. And b is on the y goes m x plus b equation, the y intercept. So that is one over v max. And so where I, again, I'm a math nerd. So I love the fact that that equation just plots really well to that graph. And you can, you don't even have to memorize the equation. The graph can get that from the graph, knowing what those points are. But just really reizing, it's the exact same stuff. They literally just rearranged the equation and then they just replotted it. No new data. It's not a whole new experiment. They didn't do the, they just took me kill us, mentors where I can just like, hey, let's make it our own kind of deal. Yeah. Yeah. Very literally. So hopefully we've won you over on line, we've reberk again, it is incredibly fast to navigate once you know what you're looking for. Rather than having to go through all these steps, you just look at x axis or x intercept, y intercept, and you're done. When we talk about inhibition, this is going to become even more important. Because what we are going to see is that these inhibitors are going to change our k m and v max in characteristic ways. Okay, we're going to talk about a couple different types of inhibitors and they each have a unique pattern of what they exactly do, what they bind to, whether it's the enzyme alone or the enzyme with the substrate, right? Where they bind on the enzyme, right? Their unique pattern of, or their unique mechanism, I should say, determines how they change k m and v max. And when you're trying to figure out what the reaction would look like, or what the line we were birth plot would look like with and without an inhibitor, the beauty is if you know that a particular type of inhibitor, let's say decreases your v max. Find the plot that has a change in the Y-intercept You're done if you know that KM stays the same Okay, find the plot that has no difference in the exit intercept It's so it's so fast and that's the beauty of it So let's get into inhibitors so we can get you hooked online. We were Burke like we are absolutely because I'm in love But you're right and so when we talk about inhibitors I think it's important to know about why we why inhibitors exist right and this kind of goes back to the idea of Regulation now we've talked about I know in previous episodes about homeostasis, right like how we the body wants to be In balance runs to be in this idea of homeostasis. We don't make do to the too hot We don't want to be too cold. We want to be just right but in order to be just right We don't want some things happening all the time So for example like we have in ourselves Millions of enzymes happening all catalyzing reactions, but maybe we actually don't want those enzymes Working all the time in all these ways and so we have all these regulatory mechanisms But one of them is by adding an inhibitor molecule now an inhibitor is what they're going to do is going to Inhibit that enzyme. That's what again the name tells you what they do and so But one day we talk about these molecules inhibiting those different ways that a molecule can inhibit them And that's what we're going to dive into and I like for separating kind of the two overarching headings right there's some inhibition That's reversible and some inhibition that is irreversible So let's talk about what we mean by reversible irreversible when we talk about something's reversible That means that inhibitor combined but because it's reversible it can come off it can let go Whereas if it's irreversible is an irreversible inhibitor when it binds It doesn't let go right. It's just stuck there and so when we talk about Irreversible inhibitor the sort of one type of inhibitor that got me I'm cat That's all like what you really need to know about it. We call it a suicide inhibitor when that inhibitor binds It binds and doesn't let go but what's important the MCAT's going to expect you to know is the bond that it makes when you see the certain bond That means you know it's not coming off and so when this inhibitor binds it binds It makes a covalent it combines covalently to the enzyme and because it's bound covalently it's not coming off and that's why we say it's irreversible And so this is important to know if that means for all of our Reversible inhibitors do any of them bind covalently? No Absolutely not if you ever see in a passage like it doesn't matter where it binds like a Irreversible inhibitor combined on the allosteric side combined at the active site if it binds Irreversible and have done it to suicide inhibitor, but that means all these reversible inhibitors the kind of the meat of today Which are reversible inhibitors they buy they have all sorts of interactions ionic, you know Hydrogen you know hydrogen bonding hydrophilic hydrophobic none of them though covalently bond is the key yes Yes, I always I always imagine it as the difference between you know, let's say the light is on You can flip the light switch and turn it off That's like an inhibitor, right? And you can just as easily flip it back on if the conditions were different a suicide inhibitor Or a irreversible inhibitor is like taking a baseball bat to the light to the light bulb is like you're done You're not taking that back. Yeah, it's not gonna turn on because you've destroyed it you fundamentally destroyed it You can't there's no going back from that you can't piece it back together and make it work, okay? So reversible inhibition is all about flipping the switch so to speak or at least a lot of them are about flipping the switch Some are actually just a major inducence Let's start our discussion off With our competitive inhibitors we we were talking about this I feel like we both kind of agree competitive is the best place to start because it's the most intuitive Most ways although I am very partial Uncompetitive is my favorite the drama is high with our uncompetitive inhibitors, but we'll start with competitive The whole idea right we have an enzyme. We know it has an active site. It's got a whole you know range of possible allosteric sites other places that things combined a competitive inhibitor is going to compete with the substrate So where is it gonna bind Obviously where the substrate is gonna bind which is the binding site or the active site? Yes exactly so competitive inhibitor just very literally like the substrate can't bind to the enzyme Because there's something else you know sitting in its spot right the competitive inhibitor is already there and Therefore the substrate and the enzyme just can't get close together. I want to be super clear The enzyme itself is not active, but it's not because the enzyme is broken. It's just because It can't get it can't grab onto the substrate and catalyze reaction Right, there's nothing fundamentally wrong with the enzyme here, right? But it can't grab the substrate right so what we see with the competitive inhibitor is usually it very closely mimics The structure of the substrate sometimes very specifically you'll hear substrate analog If you hear that you should think competitive inhibitor, right? It just means that you look the same and if you look the same You probably bind to the same spot right but as far as KM and Vmax go right what this is doing is It's preventing the substrate and the enzyme from coming together Does that sound more like something that would affect KM or Vmax? Definitely something I will initially I'm thinking it will has to do with binding It's has to do with affinity so therefore that's related to KM has to be yeah and is affinity going up or down? Oh, well, I'm thinking person foremost well. I'm less likely to bind so affinity is going down Which means KM has to be going up right there inversely related right so KM goes up Which I mean means affinity goes down. I like always kind of saying it as like I was kind of give the example of like a broccoli in a cookie Like I have a broccoli in a cookie Right I should be eating the broccoli I am the exact I have such a sweet tooth such a sweet tooth But I'm a broccoli in a cookie knowing that I have a sweet tooth I am much more likely to eat the cookie and much less likely to eat the broccoli My affinity for the broccoli has gone down my likelihood that I'm gonna eat it has gone down Therefore the KM is going to go higher. However, the enzyme itself Doesn't change yeah, and so this is important when we talk about its ability kind of like that K cat idea that we talk about It's maximum work if I don't change it Right am I gonna impact the maximum on a work that it can do? No, so therefore if I don't impact K cats, am I gonna be impacting Vmax? Nope, absolutely not so this is what we say for competitive inhibitor KM Increases because affinity decreases and Vmax does not change a great way to think about it is that we can also And this is the one that normally people learn about in school like you can't you can outcompete the competitive inhibitor Yeah, right where it's like you can add enough substrate so that you have so much substrate in relation to being inhibitor that you can just Like outcompete it at the active site and so you can still reach the same Vmax you'll just need more substrate to do so And so I I personally think about structurally especially when it comes to the other inhibitors I feel like it helps but you can think about both ways both of them equally correct where Vmax isn't gonna change Then less but KM is going to change yeah Vmax and I don't know if we said it in this this language before Vmax occurs when we say the substrate is The enzyme is saturated with the substrate meaning that's what I was alluding to and I said it's like drowning in substrate It's everywhere. It's outnumbered right there is so much substrate that the enzyme Cannot work any faster right it's overwhelmed by substrate basically at Vmax The enzyme is saturated with substrate because substrate is very high Imagine picking you know a marble out of a bag and you have Five blue marbles. That's the substrate if you had five Orange inhibitor marbles. It's gonna mess you up a lot right you're gonna pick a lot of the inhibitors by accident Again, you don't know what you're picking the enzyme can't really tell you apart doesn't have eyes right? It's just reaching out and binding whatever it can or grabbing whatever it can if we then take a million blue marbles That are the substrate and you still have five inhibitor orange marbles Are you ever gonna grab that like that? Not really no no it doesn't matter because they're so outnumbered And that's what it means to outcompete the inhibitor But okay, let's we know a competitive inhibitor KM goes up Vmax is the same What would we see in our graphs then if we we applied it there? Well, I'm just thinking about intercepts in Land River Burke So I'm just thinking okay, let's think of the am I bringing the easy one just Vmax well Vmax is not going to change Therefore my Y intercept is not going to change right one over Vmax is not it's gonna be the exact same and then okay So that's the first but I think that's like the really important point with competitive inhibitors You if you how do you know if there's a competitive inhibitor on the Land River Burke when you add the inhibitor Why just up doesn't change you're done most of the time But maybe they give you two graphs where oh the line intercepts are the same Well now I would look at my x intercept now This is where I think we're the real difficulty of Land River Burke plants because it's an inverse plot Because we have to think about this for a second because we just said a competitive inhibitor increases KM But we have to recall on the x intercept. It's one over KM and so we are Increasing the denominator therefore the number as a whole gets smaller Right and so how that would look like on the graph the K the x intercept would shift to the right Right because one over KM magnitude wise would get smaller even though the KM is getting larger and so that's how you would determine that we have a Competitive Hibber it would one over KM would shift to the right and wind us up would say it in the same Yes, yeah And again, we'll see if we could get some pictures up here. But I mean, like you mentioned, if VMAX stays the same, it's competitive. It's competitive, at least compared to the other ones. And for your information, we're going to talk about four different types of reversible inhibitors, four types, four answer options. I won't say it's a guarantee, but vast majority of the time, if you get a question about inhibition and a line we were birthplot, you're going to get four plots. One's going to be each type. Okay. So they could throw in a plot that doesn't exist. It doesn't happen quite as often because of how nice it is that there's four types of line we were birthplots with these four different types of inhibitors. So that's a competitive. Okay. The next one that is incredibly fun that you love so much. It's like I like the one's after this one. This is your favorite. Definitely. I'm going to let you have fun with our mixed and noncompetitives. I love our uncompetitive inhibitors because as someone who likes to imagine like the analogous situation in human life, because it helps me remember so much uncompetitive inhibitors are hilarious. Okay. They are so funny. And let me explain first what they do. And then I'll explain the way that I see it. In uncompetitive inhibitor and basically all the rest of the three, they're all all hysteric. They're not binding to the active site. They're binding somewhere else. Okay. They're binding to an allosteric site, but only when the enzyme and the substrate have already bound together, meaning if you have an uncompetitive inhibitor and just the enzyme, there's no inhibition. It will only inhibit when the enzyme and the substrate is together or when they are together. Right. So it will bind this allosteric site when they're together and what it does is it locks them together. It absolutely locks them together, almost glues them together or handcuffs them together. They cannot fall apart or stuck, but they also can't do anything. It also flips the theoretical switch off. I think it is so it's so funny to me because it's like, I want to catch you in the act is kind of how I see this. Right. I am not going to come on. It's almost like someone trying to catch their significant other cheating or something like I'm just going to hang out here and wait until you're together. And then I'm going to catch you in the act when you're together, snap a photo that will be you two together forever. And yet it's a photo and you can't do anything. Right. You can never be together. Right. You're memorialized in this picture or like, you know, your handcuffed together. You can never go bowling or do anything fun together. You can never create a product because you're physically tethered. You can't do anything. Right. No products can be made. Right. This does turn the enzyme off, which does mean K cat is affected. If K cat is affected, V max is affected. And specifically the speed is going down. Right. The enzyme is turned off. V max goes down, but where where the, the silliness comes in, right. Is the fact that the inhibitor is lying in weight, right. It's waiting to catch them and the act is waiting for the enzyme and substrate to already be together. And it is locking them together and cuffing, gluing however you want to think about. It is keeping them together and they cannot fall apart. Right. Traping them together, which means the K M. Well, actually, the affinity, the affinity goes up like crazy. Yeah. And this is why it's so important to talk about K M and V max is very different because the enzyme cannot work. V max and K cat are down, but because you've locked the enzyme and substrate together, the affinity is very high because the affinity is how much do you want to be together? Basically, which means K M, the nice high is also dropping. And that breaks a lot of people's brains because they're like, higher affinity is good. How could that be an inhibitor? Yeah. High affinity, sure. But if the enzyme can't do its job, what's the point? What's the point? Like there's no, there's no point to any of this. And that's why I find it to be such a fun one to think about, a fun one to explain. Both K M and V max drops. That's not a good thing because the enzyme doesn't work. It doesn't matter that you're locked together. So that one is my favorite. I think it's, it's really wonderful. And also if you are struggling to remember, uncompetitive and non-competitive, the last thing I'll say is uncompetitive unifies. I was just going to say that. It's like the unified rates brings together. Yeah. We have a, this is uncompetitive that unifies and sticks them together. There's also a non-competitive. And that made me so frustrated for so long. Right. Like literally, I get it that they're neither of them are competitive inhibitor. But why is one un and why is one non? Yeah. Yeah. I would get those mixed up. So uncompetitors or uncompetitive inhibitors unify unified, right? Unifying that enzyme and substrate together catching them in the act. I love that. That's so, I love your stories. Oh, I love your story. Wait until we talk about muscles. But yeah. And then so now we can think about, okay. So we know what it will, what we can, we'll have to K M V max. Well, if we know about those two values, less thing but linear. We're Burke, less thing about what linear. We're Burke is going to look like. And of course, the two places where we have to look is the X and Y intercept. So let's think what, what do you think would happen to those two points? So both are going down K M and V max. They're actually changing and we'll talk about this in a sec. They're changing proportional to each other. So they're both decreasing. But again, if the denominator is decreasing, the actual number is getting bigger. So they're shifting either up on the Y axis or to the left on the X. What's cool though is that it actually makes a parallel line. So the slope doesn't change. And that's important to know because when we talk about mixed, there's another option that drops K M and V max both kind of annoying, but the slope changes here. If the exact same slope, it changes them proportional to each other. So it's the same line, but just shifted to the left and up to the left. But yeah, but I like just like thinking about the like the intercepts. Like again, X intercept is going to be magnitude wise larger. Why intercept magnitude wise is going to be larger now that that can. That's the kind of thing I think that is tricky. It's like you're thinking, oh, V max is going down. There's four. So the Y and just up should go down. No, no, no, it's an inverse plot. So it actually gets higher against larger. So that's the kind of trick. Be very careful. I sometimes think like in the middle of a birth plot, whatever I think is going to happen. The opposite happens. Oh, V max. I say the same thing. Yeah, everything's opposite. Everything's opposite. So you hear V max going down. Oh, it's going to get higher. Right. Just just flip your brain that way. And that's the thing about it. But now I get to talk about what I really love. Yes. Which so we've talked to a competitive gets in the way at the active site. We've talked about one of our allosteric inhibitors binds to another place. Actually changes the structure. Our first one being an uncombed of inhibitor. But now we can talk about what I think is what that one inhibitor that I think is forgotten about so much. I I the mix inhibitor gets no love. And it makes me very sad because non-combeditive inhibitors get so much love. Right. Everyone whenever asked people about like what are the inhibitors? They say, Oh, yeah, competitive non-combeditive un-combeditive in that order. And then they're like, Yep, that's it. And I'm like, why? Because when we actually it's interesting how the MCAT does this, but in a lot of other textbooks and that I when I was learning about it, like in school, we actually framed non-combeditive inhibitors as a type of mix inhibitor, which I think is strange because the MCAT says, Oh, yeah, they're two completely different things. But therefore to understand it, we actually need to before we understand a non-combed inhibitor, we first need to understand a mix inhibitor if a non-combeditive inhibitor is a special type of one. Right. And I think there's one point to know, especially everyone's memorized probably the V max and cam, but very few can tell me why is the thing. So let's first talk about a mixed inhibitor. Right. Mix inhibitor is an allosteric inhibitor. So it doesn't bind to the active site. It binds somewhere else, but okay, how does it bind? Because we talked about an uncompetitive inhibitor, the unified inhibitor can only bind at the enzyme substrate complex, but it was just enzyme. It's not going to do it. Right. Mix inhibitors though, they're flexible. They combined either the enzyme substrate complex or just the enzyme. They're not picky that way. They're not petty and like that where they're trying to catch their spouse, trying to cheat. No, no, no, they'll bind to the enzyme, no matter what. They'll bind to it, whether the substrates there are not. So right. But the thing about mixed inhibitor is that it will have a preference. Yeah. It will prefer like again, it combined to both. But it's like, yeah, I'd rather buy the enzyme substrate complex or it might rather bind the enzyme. And this is an important idea because let's take each situation into account. Let's say it bounced the enzyme substrate complex. What do you think would happen to the affinity if it bounced the enzyme substrate complex of that specific enzyme? Well, if it prefers the enzyme substrate complex, it's going to be a little similar to our uncompetitive inhibitor. Right. It's mimicking that in its preference and it's probably going to have the same effect on the KM, which would be to drop it, drop the KM. Right. Because if it's binding to the enzyme substrate complex, it's kind of locking them in together, kind of that idea, increasing the affinity, decreasing the KM. But okay, so it would decrease the KM if it bounced the enzyme substrate complex. Right. Well, if it just bounced to just the enzyme, what do you think it would do to, in that case? So we didn't officially say this, but that's That's what a competitive inhibitor will do. Competitive inhibitor can't bind if an enzyme is already bound at substrate. So if that was its preference, it's probably going to mimic the KM patterning of competitive. If it will only bind when it binds to the enzyme alone, it's probably going to increase your KM. Right. Because it decreases. They have any kind of like the competitive. This is why we call it a mixed inhibitor. Because it's kind of a mix between an uncompetitive and a competitive. But while it can do both, it will have a preference. And so it will prefer either the enzyme substrate complex or just the enzyme. So if I asked you, Molly, you just straight up, how does a mixed inhibitor affect KM? What would you tell me? It depends. It depends what it prefers. It depends what it's. If it preferred the enzyme substrate complex, while it combined both, if it prefers the enzyme substrate complex, KM would go down. If it prefers the enzyme, it can bind both. But if it prefers the enzyme, KM will go up. Now this is the tricky thing you were talking about before, especially when it comes to the LinnuvaBerplons. Because now we have multiple options for a LinnuvaBerplons depending on our mixed inhibitor. Because before, it's always going to look like this for the other two. But now we have to take both situations into account. So let me actually, let me give you a little bit of a test. Well, I'm going to put you on the spot. I'm reading the buttons. I'm going to put you on the spot. I'm going to give you a scenario. Can you tell me whether this is a mixed inhibitor that prefers the enzyme substrate complex or if it prefers the enzyme? I think this is going to fun. So let's say I've got a LinnuvaBerplons. That's my line, KM, one over KM, one over Maxi, I just have that straight line. But then I have a new line, I have an inhibitor. Now my line, the Y intercept is now higher. It's now higher up on my Y-axis. My X intercept is now closer to zero. Okay. So the way that I would do this is think if Y-axis, if basically my Y intercept is getting further from zero and my X intercept is getting closer to zero, those are opposite patterns, which means one must be going up and one must be going down. We didn't officially say this, but mixed inhibitors, no matter what, they drop V-max. Okay. So the X-axis dropping, KM must be getting bigger, right? Which one does that resemble? It would resemble a competitive inhibitor, which means that this must bind to the enzyme alone. Right. Right. Absolutely. Because the KM is getting larger. That's why the Y intercept is getting magnitude-wise smaller. You're going to have to prep a magnitude because it's negative. But again, if I say closer to zero, it's closer to zero, right? One's further, one's closer to zero. Right. And that is an important note that I didn't touch on here, but we have to recognize that when we talked about the competitive inhibitor, we said it doesn't change the structure, so it doesn't change the V-max. All of our allosteric inhibitors bind someplace not at the active site. Yet, it's changing something at the active site. This is big, globular protein. This enzyme is binding on the other side of this, changing something at the active site. And for all allosteric inhibitors must change the structure of our enzyme. Because all the allosteric inhibitors change the structure of our enzyme, change the structure of our worker, therefore all the allosteric inhibitors must decrease V-max. Right. All of them do. And so this is something I overlook, because usually I completely forgot to mention that. And so thank you for catching that. All the allosteric inhibitors decrease V-max. It doesn't matter which one. They all do. So it makes inhibitors because their allosteric decreases V-max, but how it changes KM depends on what it prefers. And so in the other scenario that we didn't mention, you could have a scenario where both the Y intercept is now larger and the X intercept is further from zero. While that sounds like an uncompetitive inhibitor, the main difference is that the slope will be different. The slope will be different. It will prefer, again, if we talk about this mixed inhibitor, it will prefer the enzyme substrate complex kind of like an uncompetitive inhibitor. But how it changes KM and V-max are disproportionate. So therefore, the slope would be different. And so this is again where Y memorizing the graph shapes doesn't work because of the nuances that we just talked about. But there's not only one shape of a mixed inhibitor, it depends because there's variability. And so now I get to talk about one of my favorite things, which is the non-competitive inhibitor. Now, the non-competitive inhibitor, if it is a type of mixed inhibitor, a special type of mixed inhibitor, right? What do you think Molly combined to if it's a type of mixed inhibitor? It's got to be able to bind both. Right? If a mixed inhibitor combined both, interesting, but doesn't have a preference is the question. So yeah, but it has to, if it's a type, it has to have that same trait. It combined both the enzyme and the enzyme substrate complex, but you might be asking, well, well, then what's the difference between a mixed inhibitor then? Well, the key here is that it does in tabipreference. And this is a very unique type of mixed inhibitor that doesn't have a preference, it will bind them equally. Well, let's think about the impact of that, right? Because of course it's an allosteric inhibitor since it's a type of mixed inhibitor, it will decrease VMAX. We know that for certain. But now we have to ask what happens to KM? Well, if it's binding the enzyme substrate complex, what will that do to KM? KM would drop. Right? If it's binding the enzyme, what will that do to KM? It would go up. But if they're binding them both with equal affinity, what's the net effect on KM? Is nothing. No, change to KM. It is so wild to me. Everybody knows that. I want to ask, when I'm teaching a class, I'm like, okay, what happens to KM with an all-combat of inhibitor? Everyone's like, oh, KM doesn't change. Nothing happens to KM. And I ask, why? Crickets. No one knows, right? Then they say, oh, it's because it's allosteric. It doesn't impact the active side. I'm like, wait a minute. Can we just talk about an allosteric inhibitor? It did. Right? It did change KM. So it's this idea because it does. It does change KM on the micro scale. Each individual enzyme, it is changing affinity, but on the macro scale, when you take all the enzymes into account, it net does not change KM. That equal kind of idea. And I love when people realize this, they're like, oh, it makes so much sense. Why would I memorize that? And that's my question, too. This is not something to memorize. This is something to understand. But now, and that's why I like teaching it as like type of mixed inhibitor, because I want to get mixed inhibitor some love, because I think they get forgotten about. And in every biochemistry class I've ever heard or seen, and even in many common MCAT textbook series, they'll teach you competitive, non-competitive, uncompetitive. And then they bring in mixed as like, there's this other thing that's weird and doesn't even have a preference. No pattern. Don't even need to know anything about it. And it's like, we just, we memorize the three that are clear. And then there's this kind of murky fourth that no one thinks is important. But I agree flipping it, it actually makes sense. And then you can actually understand what non-competitive is, rather than just taking at face value and committing it to memory. And mixed really is a wider umbrella category of just things that can bind both. So what would the graph, this is our final type of inhibitor, what would the graph look like for non-competitive? Well, in the same way that competitive, right? Because we know there's this one value that we can really zone in, same idea with non-competitive, we know KM doesn't change. That's usually the first thought. If I'm looking for a non-competitive inhibitor, I'm looking at that x-intercept. One over KM, or negative one over KM technically, is not going to change because the KM doesn't change. The y-intercept is going to be different because v-max must change. Of course, we know that. But look at your x-intercept. That's not going to be any different. But then your y-intercept must be higher because the v-max is lower. But that's really the key. Look at your x-intercept. And if it doesn't change, you've got a non-competitive inhibitor. Yes. So to give you guys the quick rundown, if you can remember, as far as enzymes go, if you can understand what each inhibitor, like what it will bind to, is that the enzyme alone, is that the end of the enzyme is a substrate complex, and does it bind the active site or an allosteric? If you can remember that, you can then remember or learn or infer what KM and v-max will do. If you know what KM and v-max will do, you can get every line of your BIRC prop. Every line of your BIRC prop. Because you're just looking at the intercepts. And you can reason if you need to. But oftentimes the intercepts alone, even just looking what is changing. They're both changing and they're changing in the same direction. Uncompetitive. Well, if it's loop is the same, I should say. Like you can fine tune it more and more and more if you need to. But oftentimes, if you know how V-max and KM change, you can pick your answer period. You don't have to memorize the chopsticks and the swords and the- I hate that. I hate that. Because people make those up way too often. I always do. I always do. That's why I hate it. This is like, once you understand it, you can never get it wrong. If you understand what the inhibitor does, like what its mechanism is, where is it going to bind, how is it going to affect the enzyme? If you know V-max, you can infer. If you know those, you can quickly infer. Not even just like you can, it's quick to identify through line we were broke. I actually think I know a lot of us are more comfortable with Minkaylis Mentin. It actually takes a lot longer to deal with inhibition on Minkaylis Mentin. To figure out what kind of inhibitor you're dealing with, if you see like without an inhibitor and another curve with, it's actually much more difficult. It takes a lot more steps. So, go get some practice with enzyme inhibition. We've got 6,000 plus free questions in our question bank. We have even a separate area for enzymes. So definitely go get practice with that. Fundamental pastures, fundamental questions with enzymes in biochemistry. Lots and lots of practice, especially now that we've covered enzymes as a whole. That being said, we are not even close to done with enzymes. - Oh my goodness, I am so happy that we're covering enzymes now because this leads us really well into the next kind of real topic focus that we're gonna head on, 'cause what we're gonna dive into for the next few episodes is gonna be, I think one of the highest yield biochem areas, I mean, I can't think of a higher yield biochem area. - I'm gonna sit. - I mean, they're involved too, so I guess they're involved. But no, I think this is where I think a lot of people have learned this topic that the next few episodes, probably eight times, eight nine times at this point in their academic career and still don't understand it. It was just always time memorize the steps, memorize the steps, but we're gonna break them down. We're gonna break it down so that we're gonna make it understand. - What are we gonna talk about the next few episodes? - Well, the next few episodes, we are going over metabolism. - I love it. I'm so excited. - One of my favorite topics. - We've really built up to this point, right? 'Cause we've talked over to all the foundations that you need to know, Weedox chemistry, enzyme, I mean, weedox chemistry, right? - Weedox chemistry. - All these things. - All these things. - It's like a crescendo of topics. It's like now that we understand these fundamentals, we now could understand aerobic respiration, which I'm really excited for. - Yes. I love talking about all of metabolism, but I'm partial. I'm partial. Actually, I don't know. I love glycosis and I love the electron transpor chain. But I also love the Pyruvate dehydrogenase complex and this is for gas- - Is this you fangirling over aerobic respiration? - I actually don't think I can pick. I was gonna say I love ETC, but I mostly love ETC because I get to teach Redox, ETC and electrochemistry. It's a great way. - It's in a two or three hour session and that blows minds, but it's really not that hard. But we've talked about redox to electro. So I'm like, okay, will this be my favorite one? I don't think I could pick a favorite. Do you have a favorite metabolic? - I'm a bedoxidation guy to be honest. I'm not, I'm not, I'm a aerobic respiration, I know. But I'm like, I'm like a wall flower here. You're watching. You're participating too, but. - I'm kind of, I'm vibing, but I love aerobic. I love aerobic respiration, of course, but I think bedoxation is really, really cool. Just because of how much energy you can get. Anyway, I'm nerding it now. But we will talk about this in future episodes. So get excited. And I think especially the content that we cover today, yes, please review it. And I want you to realize, like this fear of linear reverb barclots that so many students have, I look at linear reverb barclots now and I am so happy. - I think. - I am so happy. 'Cause I'm like, oh, I see a linear reverb. There's nothing that can ask me that I'm not prepared for. - You already analyzed it for you. - Yeah, like you see a linear reverb. Look at the intercepts you're done. You are legitimately done. And it's like case in point. So, it's like a hug. - A hug. - I'm like, I know exactly what to expect if I see this. So again, we want everyone to have that same feeling and so practice it. And then when you start looking at the intercepts, let us know in the comments. How your practice has been going with linear reverb barclots. - Yes, absolutely. - And I want to hear about it. - Yes, absolutely. We love hearing from you guys. And good luck with all of your practice over the next couple days, or I don't know when you're listening to this. But definitely tune in for our next episode where we're gonna kind of do a broad overview of metabolism and then we're gonna zoom in on everyone's favorite process that you need to know in way too much detail, which is glycolysis. - The glycolysis. - We'll catch you in the next one, you guys. Until then, happy studying and have a great rest of your day. - Say everyone. - Bye. (upbeat music)