Mechanism of Enzyme Action

Question 1

a species inter-mediate in structure
between S and P

Answer 1

Transition State

Question 2

TRUE OR FALSE
the enzymatic rate enhancement is
approximately not equal to the ratio of
the dissociation constants of the E-S
and E- transition-state complexes, at
least when E is saturated with S

Answer 2

False (Equal)

Question 3

TRUE OR FALSE
the E must stabilize the Substrate complex, EX‡, more than it stabilizes the Transition State Complex, ES

Answer 3

False (transition state complex, substrate complex)

Question 4

ensures the favorable formation of
the ES complex

Answer 4

(triangle)Gb (Intrinsic binding)

Question 5

Gb is partially compensated by
Blank due to the binding of E
and S (TS) and by Blank
(Gd) by strain, distortion, desolvation,
and similar effects

Answer 5

entropy loss, destabilization of ES

Question 6

TRUE OR FALSE
the smaller the difference in
energies between ES and EX‡, the
Slower the E-catalyzed reaction

Answer 6

False (faster)

Question 7

Blank of the substrate
deepens the energy well of the ES
complex and actually lowers the
rate of the reaction

Answer 7

tight binding

Question 8

can involve structural strain, desolvation,
or electrostatic effects

Answer 8

destabilization of the ES complex

Question 9

a consequence of the fact that the E
is designed to bind the transition
state more strongly than the S

Answer 9

Destabilization by strain or distortion

Question 10

E bind the transition-state structure
more tightly than the Blank

Answer 10

S (or the P)

Question 11

How Destabilization of the E-S
Complex Affect Enzyme Catalysis?

Answer 11

Destabilization of the enzyme-substrate (E-S) complex enhances enzyme catalysis by preventing overly stable binding. When the E-S complex is less stable, the energy difference between it and the transition state is reduced, lowering the activation energy required for the reaction. This promotes the formation of the transition state and increases the overall reaction rate. By binding the substrate with moderate affinity, the enzyme facilitates a smoother transition to the enzyme-transition state complex, improving catalytic efficiency.

Question 11

How is it that the transition state X‡
is stabilized more than S at the E
active site?

Answer 11

The favorable interactions between the substrate (S) and the amino acid (AA) residues on the enzyme (E) contribute to the intrinsic binding energy, #Gb..

Question 12

TRUE OR FALSE
solvation of charged groups on a
substrate in solution releases
energy, making the charged
substrate more unstable

Answer 12

False (Stable)

Question 13

if the charge on the S is diminished or lost in the course of reaction, electrostatic destabilization can result
in Blank

Answer 13

rate acceleration

Question 14

when a S enters the active site,
charged groups may be forced to
interact (unfavorably) with charges of like sign, resulting in Blank

Answer 14

electrostatic destabilization

Question 15

• a “moving target”
• exists for about 10-14 to 10-13 s

Answer 15

transition state

Question 16

electrostatic destabilization of a substrate may arise from Blank of like charges in the active site

Answer 16

juxtaposition

Question 17

if such charge repulsion is relieved in the course of the reaction, electrostatic destabilization can
result in a Blank

Answer 17

rate increase

Question 18

pyrrole-2-carboxylate binds to pro racemase Blank more tightly
than L-proline, the normal S

Answer 18

160 x

Question 19

TRUE OR FALSE
dihydroxyacetone phosphate binds 40,000 times more tightly to
yeast aldolase than the substrate Phosphoglycolohydroxamate.

Answer 19

False (Phosphoglycolohydroxamate binds 40,000 times more tightly to
yeast aldolase than the substrate dihydroxyacetone phosphate.)

Question 20

Blank of purine ribonucleoside has been estimated to bind to adenosine deaminase with a KI of 3 x 10-13 M

Answer 20

1,6-hydrate

Question 21

What are the Mechanism of Catalysis ?

Answer 21

1. Near-Attack Conformations (NACs)
2. Protein Motions
3. Covalent Catalysis
4. General acid-base catalysis
5. Low-Barrier Hydrogen Bonds (LBHB)
6. Quantum Mechanical Tunneling in Electron and Proton Transfers
7. Metal ion Catalysis
8. Noncatalytic Residues

Question 22

• the reacting atoms are in van der
  Waals contact and at an angle
  resembling the bond to be formed
  in the transition state

Answer 22

Near-Attack Conformations (NACs)

Question 23

in the absence of an E, potential
reactant molecules adopt a NAC
only about Blank of the time

Answer 23

0.0001%

Question 23

NACs have been shown to form in E active sites from Blank to blank of the time

Answer 23

1% to 70%

Question 24

* proteins are constantly moving * bonds vibrate, side chains bend and rotate, backbone loops wiggle and sway, and whole domains move with respect to each other * E depend on such motions to initiate and direct catalytic events

Answer 24

Protein motions

Question 25

Protein motions may support catalysis in several ways. Active site conformation changes can (5)

Answer 25

* assist substrate binding * bring catalytic groups into position around a substrate * induce formation of a NAC * assist in bond making and bond breaking * facilitate conversion of S to P

Question 26

acceptor group on the E must be a better attacking group than Y and a better leaving group than X

Answer 26

Covalent catalysis

Question 27

formation of covalent bonds between E and S

Answer 27

BX + Y → BY + X

Question 28

enzymatic version

Answer 28

BX + Enz → E:B + X + Y → Enz + BY covalent intermediate

Question 29

readily attack electrophilic centers of S, forming covalently bonded E-S intermediates

Answer 29

nucleophilic centers for catalysis

Question 30

what are the side chains of AA in proteins?

Answer 30

✓ amines ✓ carboxylates ✓ aryl and alkyl hydroxyls ✓ imidazoles ✓ thiol groups

Question 31

* a proton is transferred in the transition state * may increase reaction rates 10- to 100-fold

Answer 31

general acid-base catalysis

Question 32

is often the most effective general acid or base because the pKa of the histidine side chain is near 7

Answer 32

histidine

Question 33

* when the barrier-to-hydrogen exchange has dropped to the point that it is at or below the zero point energy level of hydrogen

Answer 33

Low-Barrier Hydrogen Bonds (LBHB)

Question 34

the stabilization energy of LBHBs may approach Blank in the gas phase and Blank or more in solution

Answer 34

100 kJ/mol, 60 kJ/mol

Question 35

TRUE OR FALSE as the two pKa values diverge, the stabilization energy of the LBHB is increased

Answer 35

False (decreased)

Question 36

a Blank in an enzyme ground state may become an LBHB in a transient intermediate, or even in the transition state for the reaction

Answer 36

weak H bond

Question 37

* if an atom or electron is transferred in a chemical reaction from one site to another across an activation barrier, there is a finite probability that the particle will appear (as part of theP) on the other side of the energy barrier, even though it cannot achieve sufficient energy to reach the transition state

Answer 37

quantum theory

Question 38

What are the 2 Metal Ion Catalysis?

Answer 38

1. Metalloenzyme 2. Metal Activated

Question 39

* if the E binds the metal very tightly or requires the metal ion to maintain its stable, native state

Answer 39

metalloenzyme

Question 39

* E that bind metal ions more weakly, perhaps only during the catalytic cycle

Answer 39

metal activated

Question 40

is an endoprotease (it cleaves polypeptides in the middle of the chain) with a catalytic Zn2+ ion in the active site

Answer 40

Thermolysin

Question 40

Role of a metal includes act as Blank, stabilizing the increased e- density or (-) charge that can develop during rxn

Answer 40

electrophilic catalysts

Question 41

coordination to a metal ion can increase the Blank of a nucleophile w/ an ionizable proton

Answer 41

acidity

Question 42

* raising or lowering catalytic residue pKa values through electrostatic or hydrophobic interactions * orientation of catalytic residues * charge stabilization * proton transfers via hydrogen tunneling

Answer 42

Noncatalytic residues

Question 43

What are the 2 roles of metal?

Answer 43

1. Act as electrophilic catalysts, stabilizing the increased e- density or (-) charge that can develop during reaction. 2. Provide a powerful nucleophile at neutral pH.

Question 44

* a class of proteolytic enzymes whose catalytic mechanism is based on an active-site serine residue

Answer 44

Serine Proteases

Question 45

What are the 3 digestive enzymes?

Answer 45

1. Trypsin 2. Chymotrypsin 3. Elastase

Question 46

Blood-clotting enzyme

Answer 46

Thrombin

Question 46

Bacterial Protease

Answer 46

Subtilisin

Question 47

Breaks down the fibrin polymers of blood clots

Answer 47

Plasmin

Question 48

* cleaves the proenzyme plasminogen, yielding plasmin * minimizes the harmful consequences of a heart attack, if administered to a patient w/in 30 min of onset

Answer 48

tissue plasminogen activator (TPA)

Question 49

* a serine esterase and is related mechanistically to the serine proteases * degrades the neurotransmitter acetylcholine in the synaptic cleft between neurons

Answer 49

Acetylcholinesterase (not a protease)

Question 50

* cleaves peptides on the carbonyl side of the basic AAs, arg or lysine

Answer 50

trypsin

Question 51

* cleaves peptides on the carbonyl side of small, neutral residues

Answer 51

elastase

Question 52

* cleaves on the carbonyl side of aromatic residues, phe and tyr

Answer 52

chymotrypsin

Question 53

What are the 3 polar residues—— at the active site

Answer 53

His57, Asp102, Ser195

Question 54

Blank (red) is flanked by Blank (gold) and by and Blank (green)

Answer 54

His57 Asp102 Ser195

Question 55

the catalytic site is filled by a peptide segment of Blank

Answer 55

Eglin

Question 56

is actually a depression on the surface of the enzyme, with a pocket that the enzyme uses to identify the residue for which it is specific

Answer 56

the active site

Question 57

has a pocket surrounded by hydrophobic residues and large enough to accommodate an aromatic side chain

Answer 57

chymotrypsin

Question 58

the pocket in trypsin has a Blank at its bottom, facilitating the binding of positively charged Blank and Blank residues

Answer 58

negative charge (Asp189) arginine and lysine

Question 59

has a shallow pocket with bulky thr and val residues at the opening; only small, nonbulky residues can be accommodated in its pocket

Answer 59

elastase

Question 60

in the active sites of all these enzymes, the backbone of the peptide substrate is hydrogen bonded in antiparallel fashion to residues Blank to Blank and bent so that the peptide bond to be cleaved is bound close to Blank to Blank

Answer 60

215 to 219 His57 and Ser195

Question 61

the serine protease mechanism relies in part on a low-barrier hydrogen bond between Blank and Blank

Answer 61

Asp102 and His57

Question 62

* produced by mammals, fungi and higher plants * active at acidic (or sometimes neutral) pH

Answer 62

Aspartic Proteases

Question 63

Digestion of dietary protein

Answer 63

Pepsin

Question 64

Digestion of dietary protein

Answer 64

Chymosin

Question 65

Lysosomal digestion of proteins

Answer 65

Cathepsin D

Question 66

Processing of AIDS virus protein

Answer 66

HIV-protease

Question 67

Conversion of angiotensinogen to angiotensin 1; regulation of blood pressure

Answer 67

Renin

Question 68

* a hormone that stimulates smooth muscle contraction and reduces excretion of salts and fluid

Answer 68

angiotensoin

Question 69

* display a variety of S specificities, but normally they are most active in the cleavage of peptide bonds between two hydrophobic AA residues

Answer 69

aspartic proteases

Question 70

Aspartic proteases is composed of Blank to Blank AA residues

Answer 70

323 to 340

Question 71

*2 homologous domains that fold to produce a tertiary structure composed of 2 similar lobes, with approximate 2-fold symmetry

Answer 71

aspartic protease polypeptides

Question 72

each of these lobes or domains consists of Blank and Blank

Answer 72

2 β-sheets and 2 short a-helices

Question 73

the 2 domains are bridged and connected by a Blank, Blank

Answer 73

6-stranded, antiparallel β-sheet

Question 74

the active site is a deep and extended cleft, formed by the Blank and large enough to accommodate about Blank AA residues

Answer 74

2 juxtaposed domains, 7

Question 75

* causative viral agent of AIDS

Answer 75

human immunodeficiency virus (HIV-1)

Question 76

* cleaves the polyprotein products of the HIV-1 genome, producing several proteins necessary for viral growth and cellular infection * cleaves several different peptide linkages

Answer 76

HIV-1 protease

Question 77

* a remarkable viral imitation of mammalian aspartic proteases * a dimer of identical subunits that mimics the 2-lobed monomeric structure of pepsin and other aspartic proteases

Answer 77

HIV-1 protease

Question 78

HIV-1 protease cleaves between the Blank and Blank residues of the sequence Ser-Gln-Asn-Tyr-Pro-Ile-Val, which joins the Blank and Blank HIV-1 proteins

Answer 78

Tyr and Pro p17 and p24

Question 79

How many residue polypeptides that are homologous with the individual domains of the monomeric protease?

Answer 79

99

Question 80

structures determined by X-ray diffraction studies reveal that the active site of HIV-1 protease is formed at the interface of the homodimer and consists of 2 asp residues, designated Blank and Blank, one contributed by each subunit

Answer 80

Asp25 and Asp259

Question 81

1. IN EQUILIBRIUM STATE WHAT HAPPENED TO PRODUCT AND SUBSTRATE?

Answer 81

-At equilibrium in an enzyme-catalyzed reaction, the rates of the forward and reverse reactions are equal, so there’s no net change in the concentrations of substrate, product, or enzyme. The enzyme constantly binds and releases the substrate, maintaining a steady state with stable levels of substrate and product. Although molecules continue to interact, the overall concentrations remain balanced.

Question 81

2. WHY ENZYMES MUST BE DESTROYED?

Answer 81

- Their degradation helps regulate metabolic pathways, ensuring that products are neither overproduced nor depleted. Enzymes, like all proteins, have a limited lifespan and are recycled to maintain cellular health by removing damaged or misfolded ones. Environmental changes, such as shifts in pH or temperature, can also denature enzymes, preventing unwanted reactions. This natural turnover allows cells to produce fresh enzymes as needed, ensuring efficiency and proper control of cellular processes, such as growth and signaling.

Question 82

in which a substrate (S) is converted to a product (P) can be pictured as involving a transition state

Answer 82

chemical reactions

Question 83

* stable molecules that are chemically and structurally similar to the transition state

Answer 83

transition-state analogs