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## Enzyme kinetics

Current time:0:00Total duration:5:36

# Enzymatic inhibition and Lineweaver Burk plots

## Video transcript

Voiceover: So let's talk
about inhibition and how that affects enzyme kinetics. But, first, lets review
the idea that enzyme catalysis can be divided into two steps. First, the binding of enzyme to substrate, and second, the conversion
of substrate to product. And using this idea you can derive the Michaelis-Menten equation, which is useful for quantitatively looking at how enzymes behave kinetically. Also remember that within that equation, we have the Km or
Michaelis constant, which is defined as the substrate concentration where the speed of product formation is at one-half of its max value. So the first thing that
I'm going to talk about is something called a
Lineweaver-Burke plot, and how it allows us to look at the Michaelis-Menten equation
in a different way. So what I'm going to do is
take the Michaelis-Mentin equation, but then take
the inverse of both sides of the equation,
so one over everything. And that will leave us
with one over V O is equal to Km over V max times S, plus S over V max times S, since I
separated those two terms. Now if we cancel out the
two S values, then we are left with the equation
that I've just drawn out. And conveniently we can use this equation to describe a linear function. We can make one over V O
our Y or dependent variable, Km over V max our
coefficient m, or the slope, one over S our X, or
independent variable, and then one over V max our b, or Y-intercept. We can then plot this on
a graph, with our Y-axis being one over V O, and
our X-axis being one over S, and if we draw out
the corresponding line, the slope of our line will
be equal to Km over V max, and our Y-intercept will
be equal to one over V max. And we call this plot a
Lineweaver-Burke plot. And it gives us another way to look into the Michaelis-Menten equation. So let's take a step away
from this idea for a moment, and talk about the three
types of enzyme inhibitors. So our first type of inhibitor
is called the competitive inhibitor, and it works
by binding to free enzyme, or E, to form EI, or
enzyme inhibitor complex. And that will block the enzyme and make it unable to react with
substrate to form product. So in this case the
inhibitor competes with substrate for space on the enzyme. Our second type of inhibitor
is called an uncompetitive inhibitor, and it works by
binding to the enzyme-substrate complex to form ESI, which prevents the enzyme from turning
substrate into product. Our third type of inhibitor
is called a non-competitive inhibitor, which some people call a mixed inhibitor, as it can act
as both a competitive or uncompetitive
inhibitor, so it can either bind to free enzyme to
form EI, or it can bind to the enzyme-substrate
complex to form ESI, neither of which can
react to form product. So now what we're going to do is take what we just learned about inhibitors and apply it to the Lineweaver-Burke plots. So I'll start with
competitive inhibition, and I'll draw out these
three lines on the plot, labeled one, two, and three, with line one corresponding to the enzyme acting without any inhibitor around. Line two will represent
some inhibitor being present, and line three will represent even more inhibitor being present. So as you can see, as
you increase the amount of inhibitor blocking
the enzyme, the slope of these lines is increasing, while the Y-intercept isn't changing at all. What this means is that
as you increase the concentration of an
inhibitor, you're going to see an apparent increase in Km. Remember that Km is a
constant, so it's only an apparent change, and that's due to the increase in the slope of the line. But since the Y-intercept
isn't changing when you add a competitive inhibitor, you'll
see that this competitive inhibitor has no effect
on the enzyme's V max. So what that means is that
competitive inhibitors will increase Km, but leave
V max unchanged, meaning that if you really, really increase the concentration of
substrate, you'll overcome the effects of the inhibitor as you approach the enzyme's unchanged V max. But the enzyme may not be as effective at low substrate concentrations. Next we'll talk about
uncompetitive inhibition. In this case you'll notice
that all three lines have the same slope, but different
increasing Y-intercepts. What this means is that
as you increase the concentration of inhibitor, you'll see a decrease in the apparent V max, since the Y-intercept is defined as one over V max. So since you're decreasing V max as you add more inhibitor, even
if you really increase the substrate concentration,
you won't be able to overcome the effects of the inhibitor. But the enzyme will still be
effective at low substrate concentrations, even with
the inhibitor around, since there isn't that
same increase in slope. Finally, we'll take a look at non-competitive, or mixed inhibitors. And what you'll see is that this type of inhibitor, being mixed,
has characteristics of both competitive and
uncompetitive inhibitors. As you increase the
concentration of inhibitor, there is both an increase in slope and an increase in the Y-intercept. What this means is that
you'll see an apparent increase in Km, and an
apparent decrease in V max. So in this case, high
substrate concentrations won't completely overcome
this type of inhibitor, since it lowers V max,
but since there's an increase in the Km as well the enzyme will also be inhibited at low
substrate concentrations. So what did we learn? Well, first we learned
that we can rearrange the Michaelis-Menten
equation to come up with a function for the Lineweaver-Burke plots. Second we learned about
competitive, uncompetitive, and non-competitive inhibition. And third we learned
that increasing substrate concentration will only
be able to overcome the inhibitory effects of
competitive inhibitors.