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Flashcards in Enzymes pt. 1 Deck (53):
1

Enzymes are _____ specific for reactants (substrate)

highly

2

What is a holoenzyme?

An enzyme WITH a cofactor; active

3

What is an apoenzyme?

An enzyme WITHOUT a cofactor; inactive

4

What are inorganic cofactors?

Metal ions
e.g. Zn2+, Mg2+, K+, etc.

5

What are organic cofactors?

Aka coenzymes

Loosely bound, changed by reaction, derived from vitamins

6

What are the two types of organic cofactors aka conezymes?

Co-substrate

Prosthetic group

7

Describe co-substrates.

Loosely bound
Changed by reaction

8

Describe prosthetic groups

Tightly or covalently bound
Not changed by reaction

9

Can enzymes differ in their degree of specificity?

Yes

10

What is deltaG/ Gibbs free energy?

Energy of reactants - energy of products

11

Does deltaG give you any information about the rate of reaction?

No!

12

What does deltaG tell you?

If a reaction is spontaneous (G less than 0) or non-spontaneous (G greater than 0)

13

How do you calculate deltaG?

deltaG = deltaG' + RT ln ([pdts]/[reactants])

or... [C]^c[D]^d / [A}^a[B]^b

14

What is deltaG'?

Standard free energy change

15

At equilibrium, deltaG = ?

zero

16

At equilibrium, deltaG = 0, thus deltaG' = ______

deltaG' = -RT ln Keq

17

When Keq is greater than 1, _______ are favored and deltaG is ______ than zero

products, less than zero

18

When Keq is less than 1, _______ are favored and deltaG is ______ than zero

reactants, greater than zero

19

How do coupling reactions work?

They change the concentrations to make deltaG negative

20

How do enzymes lower the Ea?

Stabilizing the transition state

21

What does the transition state theory say?

The substrate and the transition species are in equilibrium

22

Keq = ?

Keq = e^(-Ea/RT)

Note, Ea also is deltaG of transition state

23

Relatively small changes in activation energy can lead to relatively ______ changes in overall reaction rate.

Large!

Because it is an exponent!

24

Once the fraction goes lower than the equilibrium fraction, free energy becomes _____ .

Negative.

e.g. if [pdt]/[reactant] = 0.5 at equilibrium, a new ratio of 0.18 is LOWER and thus deltaG is NEGATIVE

25

What are common associations/non-covalent interactions in the active site of an enzyme?

Hydrogen bonds, Van der Waals interactions

26

What is the role of reversible non-covalent bonds in enzymes?

They can be used to release free energy (bind energy)

27

What are the 2 enzyme-substrate binding models?

1. Lock and key (highly specific and rigid)
2. Induced fit (enzyme changes shape and becomes complementary to substrate)

28

How do you achieve maximum binding energy between enzymes and substrates?

Multiple short range interactions

29

Which binding model is supported by data?

Induced fit

30

How is free energy (binding energy) released?

By the formation of weak interactions from the induced fit of enzyme and substrate

31

The correct substrate for an enzyme results in more interactions and an ______ in binding energy.

Increase

Hence why substrate specificity is good

32

What are the steps of chymotrypsin deacylation?

1. Acyl-enzyme-H2O complex

2. Tetrahedral intermediate (stabilized by oxyanion hole hydrogen bonding)

3. Free enzyme

33

In addition to the use of binding energy, what other strategies do enzymes use to stabilize the transition state?

1. Covalent catalysis (covalent bond forms between substrate and enzyme)

2. Acid-base catalysis (molecule other than water becomes a proton donor/acceptor)

3. Catalysis by approximation (reactants are aligned and held close)

4. Metal ion catalysis (metal ion promotes formation of a nucleophile or an electrophile whichstabilizes the negative charge on a reaction intermediate, serving as a bridge

34

What does chymotrypsin do and where does it operate?

A serine protease that cleaves peptide bonds by hydrolysis in the gut

35

What bond does chymotrypsin cleave?

C-terminal side of aromatic (Trp, Tyr, Phe) and large hydrophobic AAs (met)

36

Is chymotrypsin hydrolysis thermodynamically favorable? kinetically favorable?

Thermodynamically favorable but NOT kinetically favorable due to partial double bond character of the peptide bond

37

What is the catalytic triad of chymotrypsin?

His - Ser - Asp

38

What is the mechanism of chymotrypsin hydrolysis?

His accepts proton form Ser hydroxyl group (acid-base)

Ser generates nucleophile to attack carbonyl

Asp stabilizes His through H bonding and electrostatics

39

What are the stages of chymotrypsin hydrolysis?

Stage 1: Acylation (acyl enzyme created)

Stage 2: Deacylation (regeneration of enzyme)

40

What are the 3 steps of chymotrypsin acylation?

1. Enzyme-substrate complex

2. Tetrahedral intermediate

3. Acyl-enzyme intermediate

41

What are the steps of chymotrypsin deacylation?

1. Acyl-enzyme-H2O complex

2. Tetrahedral intermediate (stabilized by oxyanion hole)

42

How does chymotrypsin control substrate specificity?

Binding an amino acid side chain into a deep hydrophobic cavity

43

In the acylation stage of peptide hydrolysis by chymotrypsin, His 57 accepts a proton from Ser 195. What type of catalysis does the His action represent?

General base catalysis (remember its from the perspective of histidine!)

44

Elastase is a serine protease like chymotrypsin but cleaves on the C-terminal side of residues like Ala. How would you expect the specificity pocket of elastase to compare with chymotrypsin?

Hydrophobic but smaller (alanine is relatively quite small!)

45

How do coupled reactions work?

Coupling of reactions shifts the concentration of products or reactants present and thus is able to change ΔG.

46

What is the relationship between the slope of a Lineweaver-Burk plot and catalytic efficiency under Michaelis-Menten kinetic assumptions for an enzyme?

The slope of the Lineweaver-Burk plot is Km/Vm and catalytic efficiency is defined as k3/Km. These share the term Km, but are inversely related. Vm= k3[E]t, which means that Vm and k3 are directly proportional. This means that the overall relationship between catalytic efficiency and the slope Km/Vm is an inverse one. In practice this means that steeper (numerically larger) slopes represent lower catalytic efficiency.

47

When evaluating the effectiveness of an enzyme that follows Michaelis-Menten kinetics, an effective enzyme would have:

A low Km, because this will increase the catalytic efficiency.

48

An inhibitor decreases apparent Vm but shows no decrease in apparent Km. You correctly suspect this might be:

An irreversible inhibitor or a noncompetitive inhibitor.

49

Would adding an uncompetitive inhibitor change the slope of a Lineweaver-Burk plot for an enzyme under Michaelis-Menten kinetics?

No

50

What does covalent modification do?

Add or remove charged groups to cause changes in enzyme conformation.

51

What does a positive effector do?

In allosteric control shifts equilibrium from T to R state favoring ligand binding

52

What does a negative effector do?

In allosteric control shifts equilibrium from R to T state reducing ligand binding

53

In hemoglobin(Hb) the binding of oxygen increases the affinity of Hb for oxygen. This is an example of:

Allosteric control, where oxygen is a positive effector changing Hb from the T state to the R state.