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Biochemistry > Biochem Enzymes Lecture 2 > Flashcards

Biochem Enzymes Lecture 2 Flashcards

1
Q

We study enzyme catalyzed reactions though _______

A

Enzyme Kinetics

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2
Q

In enzyme kinetics, we want to determine how ______ changes in response to experimental variables

A

Reaction rate

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3
Q

Substrate Concentration is important to rate determination, but [S] changes during the reaction as it is converted to product. We can simplify the problem by measuring the _____

A

Initial rate (initial velocity Vo)

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4
Q

What is the relationship of [S] and [E] at the start of a reaction?

So the [S] is thought of as

A

[S]»[E]

Constant

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5
Q

Vo (initial rate) can be investigated as a function of ___, which is varied by the experimenter

A

[S]

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6
Q

At low [S], Vo increases/decreases in an ~approximately linear fashion with an increase in [S].

A

Increases

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7
Q

At a higher [S], Vo increases become SMALLER/LARGER

A

Smaller until a plateau is reached

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8
Q

This is close to Vmax on the substate concentration and initial velocity curve

A

Plateau region

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9
Q

From the kinetic behavior in the graph figure, a theory was proposed

In a fast first step, the enzyme combines with its substrate to form….

In slower second step, the ES complex does what?

A

An enzyme substrate complex

Breaks down to give free enzyme and reaction product P

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10
Q

The slower reaction step that limits the overall reaction rate is called the….

Therefore, the overall rate must be proportional to the concentration of species that reacts in which step?

A

Rate Limiting Step

The slower second step

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11
Q

At low [S], the reaction rate is proportional to
[S] because the equilibrium favors formation
of ES as [S] increases

A

!

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12
Q

Vmax is reached when virtually all of the

enzyme is present as the…..

A

ES complex (no free enzyme)

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13
Q

When all of the enzyme is present as the ES complex, we can say that the enzyme is _____ with substrate. It is responsible for the plateau in the Vo vs. [S] curve

A

Saturated

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14
Q

The graph is an example of ___ kinetics. After the ES complex breaks down to P, the free enzyme can catalyze another reactant molecule

Note the a ____ state, microseconds long, occurs where ES initially builds up. The reaction quickly achieves a ___ state, where the [ES] is approximately constant

A

Saturation Kinetics

Pre-steady state

Steady State

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15
Q

E+S —> ES—-> E+P

A

Important equations

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16
Q

The measured Vo reflects the ______ state.

Analysis of initial rates are _______

A

Steady State

Steady state kinetics

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17
Q

The curve in fig. 6-11 can be expressed algebraically by which equation?

A

Michaelis-Menten Equation

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18
Q

What is the Michaelis Menten equation?

A

Vo = Vmax[S]/(Km + [S])

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19
Q

All of the terms in the Michaelis-Menten equation can be measured experimentally

A

!

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20
Q

Early in the reaction, the concentration of what is negligible?

A

P, so P—->S can be ignored

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21
Q

Vo is determined by the breakdown of ___ to ____

Therefore, Vo=?

A

[ES] to [P]

Vo = k2[ES]

22
Q

Since the concentration of ES is not measured easily, the Vo = k2[ES] equation must be expressed differently

We begin by finding what?

A

The concentration of the enzyme

[total enzyme]

23
Q

How is the total enzyme concentration expressed?

Therefore, how do you solve for free Enzyme?

Also, [S]» [Et], so the amount of substrate
bound by ES is negligible compared to [S]

A

[Et] = [E] + [ES]

[E] = [Et] –[ES]

!

24
Q

In step 1, what is the rate of ES formation?

What is the rate of ES breakdown?

A

=k1([Et] – [ES])[S]

=k-1[ES] + k2[ES]

25
Q

In step two, what is the equation representing the steady state assumption. It represents that the rate of ES formation = the rate of ES breakdown

A

k1([Et] – [ES])[S] = k-1[ES] + k2[ES]

26
Q

What is the final equation that allows you to solve for [ES]

A

[ES] = k1[Et][S]/(k1[S] + k-1 + k2)

27
Q

What is the Michaelis-menton equation?

There were other steps to get to this equation. Do you need to know?

A

Vo = Vmax [S]/(Km + [S])

28
Q

The Michaelis-Menten equation is the rate equation for an enzyme catalyzed reaction that has how many substrates?

A

One.

29
Q

When Vo=1/2Vmax, Vmax/2 =?

Therefore, we can divide by Vmax and get what equation? Important equation!

A

Vmax[S]/(Km + [S])

Km=[S] when Vo=1/2Vmax

30
Q

All enzymes that exhibit a ____ dependence of Vo on [S] follow Michaelis-Menten Kinetics

A

Hyperbolic dependence

31
Q

This is the assumption that after a very short time, the concentration of ES remains constant because ES formation = ES brekdown

A

Steady State Assumption

32
Q

Is the steady state assumption in the Michaelis-Menten equation?

A

Yes. Analysis of Vo are steady state kinetics

33
Q

When does Vmax occur?

A

When all of the E is in the form of an ES complex

34
Q

The Michaelis-Menten equation does NOT depend on
the two- step reaction mechanism proposed by
Michaelis and Menten as follows:

A

FIND THE EQUATION IN THE BOOK!

35
Q

What is the important limitation of the steady state approach

A

the meaning of Vmax and Km can be different for different enzymes

36
Q

A more convenient plot than the one in figure 6-12 for approximating Km is a double reciprocal plot, known as a ___ plot

A

Lineweaver-Burk plot

37
Q

What are the axes of the Lineweaver-Burk plot?

For enzymes which obey Michaelis-Menten kinetics, what shape is the data?

What is the slope equal to?

What is the Y intercept?

What is the X intercept?

A

1/Vo and 1/[S]

A straight line

Km/Vmax

1/Vmax

-1/Km

This linear plot gives a more accurate measure of Vmax, which is only approximated in the simple plot of Vo vs. [S]

38
Q

The Km values for some substrates and enzymes are shown in table 6-6

The data in the table shows that Km is different for different enzymes and even varies for different substrates of the same enzyme

A

For example, D-glucose and D-fructose are both
substrates for the enzyme hexokinase, but there Km
values are different (0.05 and 1.5 mM, respectively).

39
Q

A kinetic analysis gives important information about the ___ of an enzyme catalyzed reaction

For example, the substrate must form a complex with the enzyme before it can be converted to products. Complex formation is RESERVISBLE/IRREVERSIBLE

A

Mechanism

reversible

40
Q

The actual meaning of Km depends upon the number and relatives rates of the individual steps of the reaction mechanism

What is the equation for Km in reactions with 2 steps?

A

Km=(k2 + k-1)/k1

41
Q

If K2

A

k-1/k1

KD

Affinity

Smaller

42
Q

In some cases, Michaelis-Menten kinetics still apply but Km is not a measure of affinity

Sometimes k2»k-1 and Km=….

Sometimes Km remains a complex function of all 3 k values

Often, the reaction goes through multiple steps after ES formation and many rate constants define Km

A

Km= k2/k1

43
Q

In another overall reaction for Km, if the product release is rate limiting, then most of the enzyme is present in the ___ form at saturation, and Vmax = ?

____, a more general rate consant, can be used to define the limiting rate of an enzyme catalyzed reaction at saturation

A

EP form at saturation

Vmax = k3[Et]

Kcat

44
Q

In the M-M equation, what is the formula for Kcat?

Therefore, what is the equation for Vo?

A

Vmax/[Et]

Vo=kcat[Et][S]/(Km + [S])

45
Q

Kcat is a ___ order rate constant and is called the ____ number

A

first order

Turnover number

46
Q

This value is defined as the number of substrate molecules converted to product in a given time on one enzyme molecule when the enzyme is saturated with substrate

A

Kcat

47
Q

The best way to compare the turnover of different
substrates by the same enzyme or the catalytic
efficiencies of different enzymes is to compare the ratio of ____ to _____

This ratio is the

A

ratio of kcat/km

Specificity Constant

48
Q

What is the rate constant for conversion of E+ S to E+ P?

A

Kcat/Km

49
Q

When [S]«Km, then Vo =?

What order is this rate equation?

What are the units?

There is a diffusion controlled limit for this rate constant, of ___ to ____

A

Vo = (kcat/ Km) [Et][S]

Second order

M-1s-1

10^8 to 10^9

50
Q

T/F Many enzymes catalyze reactions with

multiple substrates

A

True

51
Q

In the reaction catalyzed by hexokinase:
ATP + glucose → ADP + glucose 6-phosphate
______ kinetics can be used to analyze the rates of this bisubstrate reaction

Enzymes with two substrates usually involve a transfer of an atom or ______ from one
substrate to a different one.

A

Michaelis Menten kinetics

Functional group

52
Q

What are the different pathways of enzymes with two substrates may proceed by?

A

Ternary-complex

Ping-Pong (double displacement)

Biochemistry (27 decks)

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