Chapter 9 Catalytic Strategies
Matching Questions
Use the following to answer questions 1-10:
Choose the correct answer from the list below. Not all of the answers will be used.
a) hydrolysis
b) stopped-flow
c) site-directed mutagenesis
d) chymotrypsin
e) zinc
f) stereochemical monitoring
g) magnesium
h) two-fold rotational
i) methylation
j) peptide bond cleavage
1. ____________ An enzyme that temporarily undergoes covalent catalysis as part of its
mechanism.
Ans: d
Section: 9.1
2. ____________ The type of reaction catalyzed by proteases.
Ans: a
Section: 9.1
3. ____________ The metal ion required by carbonic anhydrase for activity.
Ans: e
Section: 9.2
4. ____________ The process by which chymotrypsinogen is converted into active chymotrypsin.
Ans: j
Section: 9.1
5. ____________ A technique that permits rapid monitoring of enzyme kinetics.
Ans: b
Section: 9.1
6. ____________ The process by which host DNA is protected from cleavage by the host
restriction endonuleases.
Ans: i
Section: 9.3
Chapter 9 Catalytic Strategies
2
7. ____________ A technique that aids in determination of structure/function relationships in
enzymes.
Ans: c
Section: 9.1
8. ____________ The metal ion frequently found at active sites containing phosphate groups.
Ans: g
Section: 9.3
9. ____________ A technique that can be used to determine mechanisms when chiral molecules
are involved in reactions.
Ans: f
Section: 9.3
10. ____________ Inverted repeats in double-stranded DNA create this type of symmetry.
Ans: h
Section: 9.3
Fill in the Blank Questions
11. For a protease inhibitor to be effective, it must be _________________ for one enzyme.
Ans: specific Section: 9.1
12. The catalytic mechanism of adenylate kinase, in which the substrates are oriented to stabilize the
transition state, is called ___________________.
Ans: catalysis by approximation Section: 9.4
13. A-T base pairs are easily interrupted, as they contain only _____ hydrogen bonds versus
_______ hydrogen bonds found in G-C base pairs.
Ans: two, three Section: 9.3
14. The mechanism of chymotrypsin involves the formation of an unstable __________________ shaped intermediate that is stabilized by the oxyanion hole.
Ans: tetrahetral Section: 9.1
15. In trypsin, the specificity pocket contains a/an ______________ residue that binds to the
positive charge of the K or R residue of the substrate.
Ans: aspartyl, aspartic, or D Section: 9.1
16. The reaction center of most carbonic anhydrases is a zinc ion bound to water and
_______________ residues of the enzyme.
Ans: histidine Section: 9.2
Chapter 9 Catalytic Strategies
17. In chymotrypsin, the tetrahedral intermediate transition state is stabilized by a structural feature
referred to as the “___________________” hole.
Ans: oxyanion Section: 9.2
18. In proteases such as papain, a ___________________ residue is activated by hydrogen-bonding
to a histidine residue.
Ans: cysteine Section: 9.1
19. Enzymes that transfer a phosphoryl group from NTP to NMP are referred to as
_______________________.
Ans: nucleoside monophosphate kinases Section: 9.4
20. A characteristic feature of NMP kinases is a sequence of Gly-X-X-X-X-Gly-Lys that is referred
to as the ______________________.
Ans: P-loop Section: 9.4
Multiple Choice Questions
21. Which amino acids in chymotrypsin are found in the active site and are participants in substrate
cleavage?
A) his, ser, asp B) his, ser C) asp, lys D) lys, arg E) his, ser, arg
Ans: A Section: 9.1
22. How is specificity determined by chymotrypsin?
A)
interaction of the active site amino acids with the substrate
B)
binding of the N-terminus amino acid at the active site
C)
covalent binding of a his residue to the substrate
D)
conformational change upon binding of substrate
E)
binding of the proper amino acid into a deep pocket on the enzyme
Ans: E Section: 9.1
23. Where does cleavage of the scissile bond by chymotrypsin occur?
A)
between a his and ser amino acid
B)
on the N-terminal side of a phe or trp residue
C)
on the C-terminal side of a phe or trp residue
D)
at the N-terminal amino acid
E)
on the C-terminal side of an arg or lys amino acid
Ans: C Section: 9.1
24. Which of the following is NOT a way in which enzymes stabilize a transition state?
A)
causing the temperature of the environment to increase
B)
covalent catalysis
C)
using binding energy
D)
general acid-base catalysis
E)
catalysis by approximation
Ans: A Section: Introduction
3
Chapter 9 Catalytic Strategies
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25. What do trypsin, subtilisin, and wheat carboxypeptidase II have in common?
A)
All contain asp in the active site.
B)
All bind hydrophobic amino acids.
C)
All are synthesized in the pancreas.
D)
All contain a catalytic triad at the active site.
E)
All contain a hydrophilic substrate-binding pocket.
Ans: D Section: 9.1
26. Convergent evolution is attributed to similarities found between
A)
trypsin and elastase.
D)
chymotrypsin and trypsin.
B)
chymotrypsin and elastase.
E)
trypsin and kinase.
C)
chymotrypsin and subtilisin.
Ans: C Section: 9.1
27. If you carried out site-directed mutagenesis of subtilisin, changing serine 221 to isoleucine, what
would you expect?
A)
a large change in KM
D)
a and c
B)
a small change in KM
E)
b and c
C)
a large change in kcat
Ans: E Section: 9.1
28. The metal most commonly found at the active site of metalloproteases is
A) zinc. B) calcium. C) selenium. D) magnesium. E) sodium.
Ans: A Section: 9.1
29. Carbonic anhydrases are necessary because
A)
spontaneous hydration and dehydration of carbon dioxide occur very slowly.
B)
spontaneous hydration and dehydration of carbon dioxide are rapid, but not at speeds
necessary for biochemical processes.
C)
hydration and dehydration of carbon dioxide are coupled to other biochemical processes.
D)
a and c.
E)
b and c.
Ans: E Section: 9.2
30. Binding of a water molecule to the zinc ion induces
A)
a hydronium ion to form.
B)
a large conformation change in the binding site.
C)
ionization of a his residue, which functions as a strong nucleophile.
D)
a lowered pKa for water, which leads to formation of a zinc bound hydroxide ion.
E)
an altered KM value.
Ans: D Section: 9.2
31. Restriction endonucleases cut DNA at specific sites. How many different patterns can be formed
by a four-base sequence combination of any four bases?
A) 64 B) 256 C) 16 D) 1024 E) 4096
Ans: B Section: 9.3
Chapter 9 Catalytic Strategies
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32. Type II restriction enzymes cut
A)
double-stranded DNA, forming a 5' phosphoryl group and a 3' hydroxyl group on each
strand.
B)
single-stranded DNA, forming a 5' phosphoryl group and a 3' hydroxyl group on the
strand.
C)
double-stranded DNA, forming a 5' phosphoryl group and a 3' hydroxyl group on one
strand.
D)
double-stranded DNA, forming a 3' phosphoryl group and a 5' hydroxyl group on each
strand
E)
single-stranded DNA, forming two hydroxyl groups and loss of a phosphate group.
Ans: A Section: 9.3
33. Eco RV cleaves cognate DNA with a specificity approximately _____ times that of non-cognate
DNA.
A) 10,000 B) 10 C) 50,000 D) 1,000,000 E) 500,000
Ans: D Section: 9.3
34. Nucleotide monophosphate kinases function to
A)
transfer the phosphate from NTP to NDP.
B)
transfer the phosphate from NTP to NMP.
C)
transfer the phosphate from NMP to NDP.
D)
phosphorylate NADH.
E)
transfer the phosphate from NTP to water.
Ans: B Section: 9.4
35. Metal ion catalysis is facilitated by any of several mechanisms, including
A)
electrophilic activity, which stabilizes negative charges on an intermediate.
B)
promoting formation of nucleophiles by affecting adjacent molecules.
C)
direct binding to substrate, increasing substrate:enzyme contacts.
D)
a and c.
E)
All of the above.
Ans: E Section: Introduction
Chapter 9 Catalytic Strategies
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Short-Answer Questions
36. Complete the structure of the catalytic triad of chymotrypsin by drawing the proper structure of
the missing residue side chain the box provided. Show the proper hydrogen bonding involved in
this triad.
O
Asp
C O
CH2
H O
CH2
Ser
Ans:
His
CH2
O
Asp
CH2
C O
H N
N
H O
CH2
Ser
Section: 9.1
37. What is the challenge for a protease to facilitate hydrolysis of a peptide bond?
Ans: The peptide bond contains a carbonyl that is not very reactive; therefore, the catalytic
mechanism must employ a feature that promotes nucleophilic attack of this carbonyl
group so the peptide bond can be cleaved.
Section: 9.1
38. How can covalent modification be used to determine the mechanism of action of an enzyme?
Ans: If a particular amino acid side chain is suspected of participating in a catalytic
mechanism, covalent modification of the residue may alter it sufficiently that the enzyme
activity is altered or inhibited. However, this method is usually confirmed by other
techniques, such as site-directed mutagenesis, to rule out other possible reasons for the
loss of activity, such as conformational change.
Section: 9.1
39. Why are substrate analogs used to monitor enzyme activity?
Ans: Enzyme assays must be designed so that formation of a product is rapidly and easily
monitored. Substrates that form a colored product are easy to observe in a quantitative
manner using spectrophotometers.
Section: 9.1
40. What caused a “burst” of activity followed by a steady state reaction when chymotrypsin was
studied by stop-flow techniques?
Ans: Chymotrypsin cleaves peptide bonds in a two-step reaction, in which the first step,
formation of the acyl enzyme intermediate, is faster than the second step, hydrolysis.
Section: 9.1
Chapter 9 Catalytic Strategies
41. What supports the theory that a catalytic triad strategy is an effective means to accomplish
hydrolysis of peptides?
Ans: A number of different enzymes, including the peptidase family, some esterases, and
others, have similar mechanisms of actions. While the strategy is similar, the actual
participating amino acids differ, suggesting a mechanism commonly employed as a result
of convergent evolution.
Section: 9.1
42. What is common strategy for cysteine, metallo, and aspartyl proteases?
Ans: All employ a mechanism whereby a nucleophile is generated that attacks the carbonyl of
the peptide bond.
Section: 9.1
43. What is the common nucleophile found in cysteine, metallo, and aspartyl proteases?
Ans: The common nucleophile is water.
Section: 9.1
44. Designing drugs to inhibit enzymes is a large part of pharmaceutical research. What are some of
the enzymatic features that would be important?
Ans: The enzyme could be inhibited by interaction of a potential drug at the active site or at a
site that alters conformation or regulation of the enzyme. The structure of natural
substrates and activators, and their binding sites, would be useful features to study a new
drug design. The binding affinity and specificity would be important, and standard
enzyme assays would be used to determine the effect of the inhibitors on Kcat, KM, and
Vmax.
Section: 9.1
45. How is the bicarbonate formed when carbonic anhydrase is present?
Ans: The zinc promotes formation of a hydroxide ion, which attacks the carbon dioxide.
Section: 9.2
46. What features of carbonic anhydrase allow the rapid hydration of carbon dioxide?
Ans: Bringing the two reactants (carbon dioxide and water) facilitates the rapid reaction rate
into proximity, and the presence of a buffer system aids in proton transfer and release.
Section: 9.2
47. What mechanism is responsible for restriction endonuclease cleavage of DNA?
Ans: An activated water molecule directly attacks the phosphorous atom in a single
displacement reaction.
Section: 9.3
48. The sequence 6 bp restriction cleavage site for EcoRV is GATXXX. What is the complete
sequence of the double stranded restriction site?
Ans: GATATC
CTATAG
Section: 9.3
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Chapter 9 Catalytic Strategies
49. P-loop domains and mechanisms are common. Why?
Ans: These structures are able to undergo large changes in conformation during the binding of
NTP and its hydrolysis, thus permitting structural diversity that can account for varied
specificity.
Section: 9.3
50. Why is a metal ion required for the activity of most NTP-dependent enzymes?
Ans: The enzyme does not bind the NTP, but binds the complex consisting of the metal ionnucleotide group.
Section: 9.4
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