Exam 2 Flashcards
1
Q
3 ways to increase rate of reaction
A
- Increase temperature
- Increase substrate concentration
- add catalyst
2
Q
Active site of serine protease
A
His 57- Acid/base catalyst
Ser 195- Nuc
Asp 102- hydrogen bond to stabilize
3
Q
Chymotrypsin unique in that
A
it acts under mild conditions
Digests broad range of substrates
4
Q
Oxidoreductases
A
oxidation-reduction reactions
ex: Alcohol Dehydrogenase
GAP dehydrogenase
5
Q
Transferases
A
Transfer functional Groups
Ex: alanine aminotransferase
Hexokinase
6
Q
Hydrolases
A
Hydrolysis (cleavage by H2O)
Ex: Chymotrypsin
7
Q
Lyases
A
Group elimination, double bond formed
ex: pyruvate decarboxylase
Aldolase
8
Q
Isomerases
A
Isomerization Reactions
9
Q
Ligases
A
Bond formation coupled with ATP hydrolysis
10
Q
Multiple enzymes catalyzing the same reaction
A
isozymes
11
Q
What provides required catalytic groups when enzyme can’t
A
Cofactor
12
Q
Metal ions
A
Can exist in multiple oxidation states, type of cofactor
13
Q
Coenzymes
A
may be vitamins
- Co-substrates: enter/exit active site like substrate
- Prosthetic groups: remains in active site between reactions
14
Q
Acid-base catalysis example
A
tautomerization of ketone
15
Q
Acid-base catalysts
A
CLAGHT
Cys, Lys, Asp, Glu, His, Try
16
Q
Covalent catalysis
A
Covalent bond between catalyst and substrate during transition state formation
Ex: Decarboxylation of acetoactetate (Schiff base), makes acetone
2 part reaction, 2 energy barriers with intermediate
17
Q
Metal catalysis example
A
acetaldehyde to ethanol
18
Q
Scissile bond
A
bond cleaved by hydrolysis is positioned near Ser 195 when substrate binds to enzyme
19
Q
Why enzymes so large if few residues require?
A
Must precisely align active site
20
Q
Transition state in chymotrypsin
A
low barrier hydrogen bond, transition state stabilized (energy lowered).
Bond between Asp 102 and His 57 becomes shorter as reaction goes on
21
Q
Catalysis is effected by active site microenvironment. Explain this in hexokinase.
A
Electrostatic catalysis
nonaqueous active site allows more powerful electrostatic interactions between enzyme and substrate than aqueous solution
22
Q
How are chymotrypsin, trypsin, and elastase evolved?
A
Divergent, common ancestor
23
Q
How has subtilisin evolved?
A
Convergent: unrelated proteins have similar characteristics
24
Q
Chymotrypsin preferences
A
cleaves peptide bonds following large hydrophobic residues
25
Trypsin preference
Arginine/Lys - basic residues
26
Elastase
Cleaves peptide bonds following small hydrophobic residues (ex: alanine, glycine, valine)
27
Controlling activity: Proteolysis
Inactive chymotrypsin ---trypsin--> pi-C ---> delta C ---> alpha C
Last 2 steps by chymotrypsinogen
More active confirmation
28
Controlling activity: inhibitors
resemble substrate chemically, partial catalysis, but don't complete reaction
29
Suicide inhibitor
antithrombin- covalent binding, shut off reaction
30
The fact that the enzyme physically combines with the substrate is suggested by
hyperbolic rather than linear curve
31
unimolecular
first order, depends only on one substrate
k= s^-1
32
bimolecular
second order, depends on 2 reactants
k = M^-1 s^-1
33
What two things does velocity concentration depend on?
substrate concentration and Km
34
Km- what's going on and what does it indicate
substrate concentration when velocity is half max, indicates dissociation of ES
Shows how efficiently enzyme selects substrate and converts it to product
35
How do you get a linear Lineweaver-Burk plot
[S] >>> [E]
ES at steady state
36
Catalytic efficency
kcat/km
high kcat to maximize
Ability to convert substrate to product
units= M^-1 s^-1
37
kcat/turnover number
units = s^-1
Rate constant when enzyme is saturated with substrate
number of catalytic cycles that each active site undergoes per unit time
38
Limits to catalytic power
1. electronic rearrangements during TS
2. frequency of productive enzyme collision with substrate, max is diffusion-control limit
3. Enzyme reach perfection when rate is diffusion controlled
39
Multi-substrate reactions
Km= each substrate is different. To find it, reaction velocity measured at different concentrations of one substrate while the other substrate is present at saturating concentration
Vmax = maximum reaction velocity when both substrates are present at concentrations that saturate their binding sites on enzyme
40
Random order mechanism
doesn't matter who binds first
41
Ordered mechanism
one must bind first
42
ping pong mechanism
one binds, product released, then the other can bind
ex: transketolase
43
Multistep reaction
Ex: transketolase
intermediate: the 2 carbon fragment removed from F6P remains on enzyme while waiting for substrate
44
nonhyperbolic reactions
ex: hemoglobin
Oligomeric protein- cooperative binding, multiple active sites
Use quadratic equation
Allosteric attraction
45
irreversible inhibitor
any reagent that covalently modifies an amino acid side chain in a protein
ex: thymidylate synthases
46
Suicide substrate
enter enzyme's active site and begin to react, just as a normal substrate would, but then gets stuck in active site
ex: 5-florouracil
47
Competitive inhibitor
Reversible
Substance that directly competes with substrate for binding to enzyme's active site
Increase Km
ex: succinate dehydrogenase is inhibited by malonate
48
___ occurs when product of reaction occupies the enzyme's active site, thereby preventing the binding of additional substrate molecules
Product inhibition
49
Substrate analogues make good inhibitors, but ____ make even better inhibitors
transition state analogs
50
Is rate of product formation = ES disassociation?
No, ES complex could be going backwards (k-1)
51
noncompetitive inhibition
Km binds to site on enzyme other than active site
Km=0
Kcat/vmax are DECREASED
ex: metal ions
52
Mixed inhibition
apparent Km may increase or decrease
Vmax decreases
53
Uncompetitive inhibition
multi-substrate reactor. inhibitor binds are one substrate has bound.
Vmax/Km are lowered to same degree, parallel lines
54
Allosteric inhibition in glycolysis: phosphoenolpryruvate.
| Why inhibit primary regulatory step in glycolysis.
To prevent unneeded energy expression.
If pyruvate accumulates, too much glycolysis, not full use of product.
Feedback inhibitor and Negative effector
55
How does phosphoenolpyruvate inhibit glycolysis?
Swaps Arg 162 (+ charge) with Glu 161 (- charge)
| F6P (- charge) no longer attracted to a + charge, is repelled instead
56
ADP
positive effector
binds to same sight as phosphofructokinase
forces ARG 162 to remain where ti can stabilize F6P binding.
57
4 factors that influence enzymatic activity
1. change in rate of enzyme's synthesis
2. change in subcellular location
3. ionic "signal"- change in pH, release of calcium
4. covalent modification
58
Most modern drugs are _____. Ideal drugs are
inhibitors
high affinity, high selectivity, non-toxic
59
As Km increases
affinity of enzyme for substrate decreases
60
During inhibition if vmax isn't 0 (as in competitive), it
Decreases
61
prosthetic group
cofactors that never leave molecule
62
What is prosthetic group in hemoglobin?
heme- iron in hydrophobic pocket between E and F.
63
6 bonds on iron
4 with nitrogen in ring
1 with histidine residue in F helix
1 with oxygen
64
Why must heme be part of a protein to be a good oxygen carrier?
by itself, central atom Fe(II) can easily be oxidized to FE(III), which can't bind O2.
65
Myoglobin and hemoglobin: structural similarities
heme group in hydrophobic pocket
His F8 on iron
His E7 H-bond to O2
HOWEVER: primary linear structure different
66
Invariant residues
essential for function. If you change them, alter molecule function (similar in myoglobin and alpha/beta subunits)
67
Myoglobin and Hb are homologous proteins meaning
common ancestor (oxygen binding proteins)
68
Conservatively substituted residues
position under less pressure to maintain particular amino acid, can be substituted by similar amino acid
69
Variable position
position that can accommodate variety of residues, none critical for structure/function
70
Deoxy
```
Large cavity
T
P. ring is bowed
low O2 affinity
Heme Fe has 5 ligands
```
Histidine residue on beta between proline and threonine residues on alpha
71
Oxy
```
small cavity
R
heme group planar
high O2 affinity
Heme Fe has 6 ligands
```
Histidine residue on beta between threonine residues on alpha
72
Bohr Effect
reduction of hemoglobin's O2 binding affinity when pH DECREASES ( [H+] increases)
73
BPG
stabilizes deoxy confirmation of hemoglobin to unload oxygen. Without, hemoglobin would bind too tightly.
5- charges that interact with + charge on hemoglobin
only binds to central cavity in T state
74
Fetus: H21 ---> Serine, so...
+ charge that interacts with BPG is gone
| reduces fetal binding, helps transfer O2 from maternal circulation to fetus
75
Microfilament functions
```
support PM
determines cell shape
structural support
cell movement
tensile strength
```
76
Actin polymer
polymerized is F-actin
+ end grows faster
ATP is - end
G-actin is globular
77
Prevent actin growth
adding capping protein.
78
Microtubule functions
reinforce cytoskeleton
construct cilia/flagella
align and separate chromosomes, form spindle apparatus
79
Microtubules structure
Binds to GTP/GDP
1 nucleotide binding site/tubulin subunit
Alpha GTP binding site buried in interface while beta exposed, GTP hydrolyzed, but resulting GDP remains bound and can't diffuse
80
Single strand microtubule called
protofilament
81
Positive end of microtubule called
beta (alpha is negative, often anchored)
82
Paciltaxel
binds to beta tubulin
| blocks depolymerization, stabilizes
83
Colchicine
destabilizes protofilament, interferes with side/side interactions.
Binds at interface between alpha/beta dimer
shut off cell division
84
Intermediation filaments: Keratin
coil-coiled.
dimer of alpha helices.
1/4th residues hydrophobic and holds coil together
7 residue repeat units
85
Keratin tetramers are antiparallel staggering held together by
cystine residues
86
Intermediate filaments: Collagen
Triple helix
every 3rd is glycine --> secondary structures
30% proline/hydroxyproline
Gly-Pro-Hyp triplet (left- more stable)
3 polypeptides in right hand triplet, stabilized by H bonding.
Staggered parallel
87
Where is collagen assembled?
| Collagen is modified/cross linked by...
Endoplasmic Reticulum
covalent modifications
88
Myosin
microfilaments
change triggered by hydrolysis of ATP bound to head
unidirectional
alpha helix lever
89
Kinesin
microtubules
moves toward + end
unidirectional
90
Why is kinesin, unlike myosin, processive or highly processive?
kinesin constantly holds microtubules, many cycles of ATP hydrolysis and kinesin advancement occur before the motor dissociates from its microtubule tract.
Myosin dissociates after 1 stroke.
