Exam 3 Flashcards
1
Q
Why biocatalysts over inorganic catalysts
A
greater reaction specificity, milder conditions, greater regulation, faster reaction rates
2
Q
The — of enzymes are essential to their catalytic
activity
A
structure
3
Q
What are some common inorganic co-factors?
A
Fe,Mg,Mn,Zn, complex organic/metalloorganic molecules
4
Q
why are co-factors important?
A
Bind to enzyme to form catalytically active enzyme
5
Q
what reaction do oxidoreductases catalyze?
A
Oxidation reduction reactions (movement of electrons)
6
Q
What reactions do transferases catalyze?
A
transfer of functional groups
7
Q
What reactions do hydrolases catalyze?
A
Hydrolysis (transfer of functional groups to water)
8
Q
What reactions do lyases catalyze?
A
cleavage of c-c,c-o,c-n & other bonds by elimination resulting in double bond
9
Q
What reactions do isomerases catalyze?
A
Transfer of atom group within the molecule to yield isomers
10
Q
What reactions do Ligases catalyze?
A
Formation of c-c,c-s,c-o, and c-n bonds by condensation
11
Q
Each enzyme has a — number classification
A
4 part
12
Q
What is the first number of the enzyme classification?
A
the class name
13
Q
What is number two of enzyme classification?
A
subclass
14
Q
What are the 3rd and 4th number of the enzyme classification?
A
provide more information about the reaction
15
Q
A specific substrate interacts with a specific group of —– in the enzyme at the active site
A
amino acid residues
16
Q
The substrate binds to an enzyme to produce?
A
The products
17
Q
Active site
A
location on enzyme in which the enzyme catalyzed reaction occurs
18
Q
Substrate
A
molecule which binds to the enzyme active site and is acted upon by the enzyme
19
Q
Enzymes Affect
A
reaction rates no the equilibria
20
Q
——– change in gibbs free energy is favored
A
negative
21
Q
A higher activation energy means
A
slower reaction
22
Q
Catalysts increase
reaction rates by
A
lowering change in gibbs free energy
23
Q
Rate-limiting step
A
Step in enzymatic reaction which has the largest activation energy barrier. Thus, dictates overall rate of reaction
24
Q
What is the reaction rate is determined by
A
the concentration of reactant and a rate constant k
25
Equation for Keq
keq=[products]/[reactants]
26
If ΔG° < 0, then
Keq>1 and products are favored at equilibrium
27
If ΔG° > 0, then
Keq<1 and reactants are favored at equilibrium
28
What helps to lower activation energies?
Weak non-covalent interactions between the substrate and enzyme
29
What are the weak Interactions between E and S in ES complex?
Hydrogen bonding, hydrophobic interactions, and ionic interactions
30
Some of the weak interactions in the ES complex occur during the reaction
transition state and
stabilize the transition state
31
binding energy, ΔGB
energy concerning enzyme and substrate interactions
32
The binding energy can be used to lower substrate entropy or
to cause a
conformational change in the enzyme (induced fit)
33
Enzymes binds to the ----- better than the -----
transition states, substrates
34
Stronger/additional interactions -----------
compared to the ground state lowers the activation barrier
between the enzyme and transition state
35
Why are enzymes large?
maximizes number of weak interactions between enzyme and transition states
36
Specificity
enzyme is selective for a particular substrate or group of substrates
37
induced fit
enzyme undergoes conformational change upon binding to substrate. Caused by weak interactions between substrate and enzyme
38
Binding energy holds substrates in the proper orientation to react resulting in
entropy reduction
39
Enzyme-substrate interactions replace all
hydrogen bonding between substrate and water
40
Catalytic residues
functional groups directly involved in a catalytic reaction
41
When there is no catalyst present, unstable, charged
intermediates formed in many biochemical reactions tend
to break down into the reactants with zero product formation
42
These charged intermediates can be stabilized by
proton transfers and be broken down into products
43
Specific acid-base catalysis
only utilizes the H+ or OH- ions present in water
44
General acid-base catalysis
proton transfer facilitated by other molecules
45
In an enzyme’s active site, amino acid side-chains can act as
proton acceptors or donors
46
Glu, Asp general proton donor form
R-COOH
47
Glu,Asp general proton acceptor form
R-COO(-)
48
Lys,Arg general proton donor form
R-NH3(+)
49
Lys, Arg general proton acceptor form
R-NH2
50
Cys genral proton donor form
R-SH
51
Cys general proton acceptor form
R-S(-)
52
Ser general donor form
R-OH
53
Ser general proton acceptor
R-O(-)
54
What is the purpose of catalysis?
Stabilize the charged intermediate to promote the formation of a product
55
What are the two major steps of covalent catalysis?
1) covalent bonding between amino acid residues in active site of enzyme and substrate
2) Cleavage of covalent bond between substrate and enzyme to return enzyme to original state
56
Covalent catalysis requires a --- and ---
nucleophile and electrophile
57
In covalent catalysis, the ---- attacks the ----
nucleophile attacks the electrophile
58
In covalent catalysis, the nucleophile may be
a reactive serine, amine, thiolate, carboxylate
59
Metal Ion Catalysis involves
ionic interaction between enzyme with metal cofactor and a substrate
60
Ionic interactions between an enzyme-bound metal and a
substrate can
orient the substrate into the correct position for reaction, or stabilize the transition state
61
enzyme bound metal participate in ---- reactions
redox
62
chymotrypsin uses -- and --- catalysis
covalent and general acid base catalysis
63
enzyme kinetics
study of how enzyme catalyzed reaction rate responds to changes in experimental parameters
64
the Michaelis-Menten
equation
V0=Vmax[S]/[S]+Km
65
Km equation
Km=[E][S]/[ES]
66
What is the definition of Km?
When the substrate concentration is equal to 1/2 of max rate. Thus, 50% of substrate is bound to enzyme
67
Which step is the fast step in enzyme action?
The binding of the substrate to the enzyme to form complex
68
Which step is the slow step(rate limiting) in enzyme action ?
Enzyme-substrate complex breaks down to generate free enzyme and product
69
The overall reaction rate must be
proportional to
The concentration of enzyme-substrate complex
70
The saturation effect
A plateau in velocity at high [S]
71
At low [S], KM>> [S], most of the
enzyme is in the free form. Thus, ---
change in velocity is linear and dependent on [S]
72
At high [S], KM<< [S]the enzyme is
saturated with substrate; almost all
enzymes are in the combined form. Thus,
change in velocity is not linear or dependent on [S]
73
steady state
[ES] and other intermediates remain approximately constant over time
74
what does steady state imply about [ES] formation and breakdown
[ES] formation = [ES] breakdown
75
The Michaelis-Menten equation can be algebraically
transformed into a more useful form in plotting
experimental data, simply by
taking the reciprocal of both sides to yield a linear equation
76
What is the slope of the Lineweaver-Burk equation
Km/Vmax
77
What is the y-intercept of the Lineweaver-Burk equation?
1/Vmax
78
Lineweaver-Burk equation
1/V=(Km/Vmax)+(1/Vmax)
79
Why is Lineweaver-Burk equation preferred over the the Michaelis-Menten
equation?
It provides a more accurate measure of Vmax
80
For a reaction with two steps, Km
=
Km=(K2+K(-1))/k1
81
What is the mathematical measure of Kd?
Kd=(k-1)/K1
82
What is Kd an indicator of?
Affinity between the substrate and enzyme
83
When is Km equal to Kd the dissociation constant?
When K2
84
if product release is rate-limiting, then most of the enzyme
is present in the --- form at saturation
EP
85
What is Kcat the rate constant for?
The rate limiting step
86
when product release is the rate limiting step what does v max equal
Vmax=Kcat[Etotal]
87
kcat is also known as
the turnover number
88
What is the turnover number?
number of substrate molecules converted to product in a given period of time
89
the overall rate constant for conversion of E+S ➔ E+P
kcat/km and measures how efficiently an enzyme turns a substrate into product
90
Enzymes with two substrates usually involve a transfer of
an atom or functional group from one substrate to another
91
Enzyme inhibitors
interfere with catalysis to slow or halt enzyme catalyzed reactions
92
Reversible Inhibition
noncovalent binding of inhibitor to enzyme . can be reversed by removal of inhibitor
93
What are the three kinds of reversible inhibition?
Competitive, noncompetitive, mixed inhibition
94
Irreversible Inhibition
covalently binds to enzyme and destroys essential functional group or portion of enzyme
95
A competitive inhibitor molecule competes with the substrate molecule
for
the active site of an enzyme
96
Usually, a competitive inhibitor is very similar to
the substrate
97
Formation of an EI complex consumes [E] thus
decreases formation of ES complex
98
In the presence of a competitive inhibitor, how does the km change
it increases and is now αKm
99
Due to reversibility of competitive inhibition, the ES complex is favored
over EI
at high [S] thus v max stays the same
100
Why does km increase in the presence of a competitive inhibitor?
a larger [S] is required to reach 1/2 v max
101
An uncompetitive inhibitor binds at a
site distinct from the active site
102
How does uncompetitive inhibition impact the V max and Km
it decreases them by a factor of alpha'
103
A mixed inhibitor binds at
a site distinct from the active site but may bind to enzyme of ES
104
When α=α’
, it is known as
noncompetitive inhibition and Km is not affected
105
Suicide inactivators are a special type of
irreversible inhibitor
106
Suicide inhibitors are inactive until
they covalently bind to the active site of a particular enzyme
107
Chymotrypsin catalyzes
the hydrolytic cleavage of peptide bonds adjacent to large aromatic amino acid residues
108
What amino acids does chymotrypsin cleave at?
Trp, Phe, Tyr and on the c side
109
What is the substrate of chymotrypsin?
a protein or peptide with Trp,Phe, and Tyr residues
110
What is the Acylation phase of the chymotrypsin mechanism
Cleavage of peptide bond to establish formation of ester linkage between carbonyl carbon of amino acid residue and enzyme
111
What us the deacylation phase of the chymotrypsin mechanism?
Hydrolysis of the ester linkage and regeneration on non acylated enzyme
112
Where is the active site of chymotrypsin?
A hydrophobic pocket next to the catalytic triad
113
What amino acid residues compose the catalytic triad of chymotrypsin?
His57,Asp102,Ser195
114
What family does chymotrypsin belong to?
The Serine protease family
115
What position is the gly residue located at in chymotrypsin?
G193
116
The acylation and deacylation of the chymotrypsin involves which enzyme?
The Serine residue
117
Which step is faster acylation or deacylation?
acylation
118
What is the rate determining step of the chymotrypsin mechanism?
The deacylation
119
During acylation, ---- as a general acid and --- as a general base facilitate the
nucleophilic attack of -----
Asp102,His57,Ser195
120
During acylation,------ acts as a general base, which can remove the proton from the ------
His57,Ser 195
121
Why is the catalytic triad important?
Aids in the stabilization of intermediates formed during the acylation and deacylation steps. Thus, lowering energy of activation
122
An optical rate of chymotrypin is observed at a pH of
8
123
Why is chymotrypsin reaction rate not ideal at low pH?
The His57 residue is pronated and unable to remove the proton from the Ser195 residue
124
Why does reaction rate decrease after pH of 8?
Deprotonation of N-terminus in one of three polypeptide chains. Hydrophobic pocket at active site closes and substrate binding decreases
125
What is the first step of Chymotrypsin mechanism?
Enzyme substrate binding
126
What is the second step of chymotrypsin mechanism?
Nucleophilic attack by ser 195 to carbonyl of substrate. Forming a tetrahedral intermediate
127
How does His 57 make Ser195 a better nucleophile?
Removing the proton from Ser residue's hydroxyl group
128
What is the third step of chymotrypsin mechanism?
The re-instatement of the carbonyl double bond resulting in cleavage of peptide bond in substrate.
129
What is step four of the chymotrypsin mechanism?
A water molecule is deprotonated by His 57 to generate a nucleophilic -OH molecules
130
What is the fifth step in chymotrypsin mechanism?
The OH nucleophile attacks the acyl enzyme ester linkage to form a second tetrahedral intermediate
131
What is the fifth step in chymotrypsin mechanism?
The OH nucleophile attacks the acyl enzyme ester linkage to form a second tetrahedral intermediate
132
What is step six of the chymotrypsin mechanism?
The tetrahedral intermediate collapses to form carboxylate anion. Ser 195 is displaced in collapse of intermediate
133
What is the final step of the chymotrypsin mechanism?
The final product diffuses away from the enzyme's active site to produce a free enzyme
134
How does inhibition of chymotrypsin occur?
Metal ion binding to the His 57
135
What serves to inhibit enzyme activity of papain
Metal ion binding to the Cys25 residue whichi is similar to the -oH group of the Ser 195 in chymotrypsin
136
Di-isopropylfluorophosphate
(DIFP) is an ---
for chymotrypsin
irreversible inhibitor
137
What amino acid reside does the DIFP inhibitor bind to in chymotrypsin?
Forms covalent bond to the Ser195
138
Transition-state analogs are molecules
that resemble
transition states
139
Binding to an allosteric regulatory enzyme affects the -- and -- structure of a protein
tertiary and quatenary
140
How does an allosteric enzyme bind to a substrate? What does the binding result in
Reversible non-covalent bonds resulting in conformational change. Conformational change results in change of activity
141
the regulatory enzyme is homotropic when
the enzyme and substrate are similar
142
ATCase is an alloesteric enzyme that catalyzes the formation of
N-carbamoylaspartate from carbamoyla phosphate and aspartate
143
What is the quaternary structure of ATcase?
6 catalytic subunits and 6 regulatory subunits
144
-- is an inhibitor of ATCase
CTP
145
Do allosteric enzymes follow Michaelis-Menten kinetics?
No
146
UMP can be converted to
UTP by ATP
147
UTP can be reacted with --- to give --
Glutamine, CTP
148
ATP and CTP are --- of ATCase
heterotropic regulators
149
n feedback
inhibition, a ------ in a sequence
of reactions inhibits
the synthesis of ----------
final product, additional product
150
The attachment of phosphoryl
groups (-PO3
2-
) to specific amino
acid residues of a protein is
catalyzed by
protein kinases
151
removal of a phosphoryl
group is catalyzed by
protein phosphotases
152
------- the most common type of regulatory modification
phosphorylation
153
The phosphoryl group comes from
Atp
154
An oxygen atom from the phosphoryl group may form a ----- with one or multiple groups of the protein
hydrogen bond repeling charged amino acid residues. This disrupts protein tertiary structure
155
To become an active enzyme -----must occur in chymotrypsinogen and trypsinogen
proteolytic cleavage
156
Hydrolysis of the peptide bond between --- and ---
Arg15 and Ile16
157
hydrolysis is followed by --- to produce three active chymotrypsin peptide chains
Lys6, Ile7
158
Hexokinase reaction
ATP + D-glucose ➔ADP + D-glucose-6-phosphate
159
In the hexokinase reaction, what must ATP be bound to?
Mg2+
160
Hexokinase undergo --- upon binding of
glucose, which allows correct positioning of active site residues for catalysis
Induced fit
161
Enolase is a glycolytic enzyme (dimer,
Mr 93,316). It catalyzes reversible
dehydration of 2-
2-phosphoglycerate to phosphoenolpyruvate
162
In the enolase reaction, ---- as as the base catalyst by removing a proton at C-2
Lys345
163
In the enolase reaction, --- acts as a acid catalyst by donating a proton to the -OH leaving group
Glu211
