1.11.1.5: cytochrome-c peroxidase

This is an abbreviated version!
For detailed information about cytochrome-c peroxidase, go to the full flat file.

Word Map on EC 1.11.1.5

Reaction

2 ferrocytochrome c +

H2O2
=
2 ferricytochrome c
+ 2 H2O

Synonyms

apocytochrome c peroxidase, BCcP, CCP, CCP1, CcpA, Cjj0382, CytC, cytochrome c iso-1, cytochrome c peroxidase, cytochrome c-551 peroxidase, cytochrome c-H2O oxidoreductase, cytochrome peroxidase, di-heme cytochrome c peroxidase, diheme cytochrome c peroxidase, diheme cytochrome c5 peroxidase CcpA, DocA, LmP, MacA, mesocytochrome c peroxidase azide, mesocytochrome c peroxidase cyanate, mesocytochrome c peroxidase cyanide, peroxidase, cytochrome c, Psa CcP

ECTree

     1 Oxidoreductases
         1.11 Acting on a peroxide as acceptor
             1.11.1 Peroxidases
                1.11.1.5 cytochrome-c peroxidase

Engineering

Engineering on EC 1.11.1.5 - cytochrome-c peroxidase

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PROTEIN VARIANTS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
W191F
-
less efficient at catalytic turnover than the wild-type enzyme
W191G
-
the mutant exhibits a loop-gated artificial protein cavity
W51F
-
exhibits extensive dimerization
A124K/K128A
site-directed mutagenesis, no significant changes
G94K/K97Q/R100I
site-directed mutagenesis, triple point mutant is created to mimic the critical loop region of, but its crystal structure reveals that the inactive, bishistidinyl-coordinated form of the active-site heme group is retained
H93G
site-directed mutagenesis
M297H
site-directed mutagenesis
S134P
site-directed mutagenesis, distortion of the loop region, accompanied by an opening of the active-site loop, leaving the enzyme in a constitutively active state
S134P/V135K
site-directed mutagenesis, distortion of the loop region, accompanied by an opening of the active-site loop, leaving the enzyme in a constitutively active state
H93G
-
site-directed mutagenesis
-
M297H
-
site-directed mutagenesis
-
F81W
-
modest changes in in vitro peroxidase assays
D37E/P44D/V45D
-
redesign of a manganese-binding site, ratio kcat/KM values for manganese oxidation is 0.33 per mM and s at pH 5.0
D37E/V45E/H181E
-
redesign of a manganese-binding site, ratio kcat/KM values for manganese oxidation is 0.25 per mM and s at pH 5.0
G41E/V45E/H181D
-
redesign of a manganese-binding site, ratio kcat/KM values for manganese oxidation is 0.10 per mM and s at pH 5.0
G41E/V45E/W51F/H181D/W191F
-
redesign of a manganese-binding site, ratio kcat/KM values for manganese oxidation is 0.6 per mM and s at pH 5.0
H71G/W94A
E117H
-
no enzymatic activity
E117K
-
no enzymatic activity
E117L
-
no enzymatic activity
H74M
-
no enzymatic activity, reduced redox potential. The introduced methionine does not ligate the N-terminal heme
M118H
-
no enzymatic activity
M118L
-
7.3% of wild-type activity
M278H
-
no enzymatic activity, reduced redox potential. Mutant contains two low-potential hemes
Q107L
-
no enzymatic activity
W97A
-
no enzymatic activity. W97 is the mediator of intramolecular electron transfer of the enzyme
W97F
-
no enzymatic activity. W97 is the mediator of intramolecular electron transfer of the enzyme
A193F
-
surface mutant, shift in reduction potential to -170 mV. Analysis of spectroscopic properties
A193W
-
mutant designed to incorporate a Trp-based extension to move oxidizing equivalents from the heme to the protein surface. Mutant is able to oxidize veratryl alcohol substrate with turnover numbers greater than wild type
A193W/Y229W
-
mutant designed to incorporate a Trp-based extension to move oxidizing equivalents from the heme to the protein surface. Mutant is able to oxidize veratryl alcohol substrate with turnover numbers greater than wild type, possibly using an electron hopping mechanism
D146N
-
surface mutant, shift in reduction potential to -173 mV. Analysis of spectroscopic properties
D146N/D148N
-
surface mutant, shift in reduction potential to -173 mV. Analysis of spectroscopic properties
D18K
-
positive-to-negative charge-reversal mutant
D210K
-
positive-to-negative charge-reversal mutant
D235A
-
proximal pocket mutant, shift in reduction potential to -78 mV. Analysis of spectroscopic properties
D235E
-
proximal pocket mutant, shift in reduction potential to -113 mV. Analysis of spectroscopic properties
D235N
D33K
-
positive-to-negative charge-reversal mutant
D34K
-
the mutation causes large increases in the Michaelis constant indicating a reduced affinity for cytochrome c
D34N
-
surface mutant, shift in reduction potential to -175 mV. Analysis of spectroscopic properties
E118K
E17K
-
positive-to-negative charge-reversal mutant
E201K
-
positive-to-negative charge-reversal mutant
E209K
-
positive-to-negative charge-reversal mutant
E290C
-
formation of a covalent complex with cytochrome c mutant K79C, kinetic studies. Residual activity of complex is due to unreacted enzyme that copurifies with the complex. In the complex, the Pelletier-Kraut site is blocked which results in zero catalytic activity
E290K
E290N
-
surface mutant, shift in reduction potential to -177 mV. Analysis of spectroscopic properties
E291K
-
positive-to-negative charge-reversal mutant
E291Q
-
surface mutant, shift in reduction potential to -162 mV. Analysis of spectroscopic properties
E32K
-
positive-to-negative charge-reversal mutant
E32Q
-
surface mutant, shift in reduction potential to -168 mV. Analysis of spectroscopic properties
E35K
-
positive-to-negative charge-reversal mutant
E98K
-
positive-to-negative charge-reversal mutant
H52D
-
distal pocket mutant, shift in reduction potential to -221 mV. Analysis of spectroscopic properties
H52E
-
distal pocket mutant, reduction potential -183 mV, comparable to wild-type
H52K
-
distal pocket mutant, shift in reduction potential to -157 mV. Analysis of spectroscopic properties
H52L |
-
site-directed mutagenesis, a distal pocket mutant
H52L/W191F
-
proximal pocket mutant, shift in reduction potential to -151 mV. Analysis of spectroscopic properties
H52N |
-
distal pocket mutant, shift in reduction potential to -259 mV, most negative reduction potential of all mutants analyzed. Analysis of spectroscopic properties
H52Q
-
distal pocket mutant, shift in reduction potential to -224 mV. Analysis of spectroscopic properties
H52Q |
-
site-directed mutagenesis, a distal pocket mutant
K12C
-
characterization of complex with yeast cytochrome c mutant K79C. Cytochrome c is covalently bound and located 90° from its primary binding site. Catalytic activity is similar to wild-type cytochrome c peroxidase
K149D
-
positive-to-negative charge-reversal mutant
K264C
-
characterization of complex with yeast cytochrome c mutant K79C. Cytochrome c is covalently bound and located 90° from its primary binding site. Catalytic activity is similar to wild-type cytochrome c peroxidase
N184R
the N184R variant introduces potential hydrogen bonding interactions for ascorbate binding
N184R/W191F
site-directed mutagenesis
N78C
-
characterization of complex with yeast cytochrome c mutant K79C. Cytochrome c is covalently bound and located 90° from its primary binding site. Catalytic activity is similar to wild-type cytochrome c peroxidase
R31E
-
positive-to-negative charge-reversal mutant
R48A/W51A/H52A
R48E
-
distal pocket mutant, shift in reduction potential to -179 mV. Analysis of spectroscopic properties
R48L/W51L/H52L
R48L/W51L/H52L |
-
site-directed mutagenesis, a distal pocket mutant
R48V/W51V/H52V
V197C/C128A
-
as active as the wild-type enzyme. Used to generate a covalent complex with a mutant cytochrome c
V5C
-
characterization of complex with yeast cytochrome c mutant K79C. Cytochrome c is covalently bound via disulfide formation of the mutated residues and located on the back-side of the enzyme, 180° from its primary binding site. Catalytic activity is similar to wild-type cytochrome c peroxidase. Significant electrostatic repulsion of the two cytochrome c molecules bound in an 2:1 complex which decreases as the ionic strength of buffer increases
W191F
W191G
provides a specific site near heme from which substrates might be oxidized
W51H/H52L
W51H/H52W
-
altered electronic absorption spectra, indicating that the heme group in the mutants is six-coordinate rather than five-coordinate as it is in wild-type cytochrome c peroxidase, weaker effect on cyanide binding, with the cyanide affinity only 2-8times weaker than for cytochrome c peroxidase
Y229W
-
mutant designed to incorporate a Trp-based extension to move oxidizing equivalents from the heme to the protein surface. Mutant is able to oxidize veratryl alcohol substrate with turnover numbers greater than wild type
Y36A
site-directed mutagenesis, Tyr36 directly blocks the equivalent ascorbate binding site in CcP and was therefore replaced with a less bulky residue
Y36A/N184R
site-directed mutagenesis, no significant spectroscopic changes on reaction with stoichiometric or higher amounts of H2O2 are seen
Y36A/N184R/W191F
site-directed mutagenesis, cytochrome c peroxidase enzyme can duplicate the substrate binding properties of ascorbate peroxidase through the introduction of relatively modest structural changes at Tyr36 and Asn184, no evidence for a porphyrin pi-cation radical
Y36A/W191F
site-directed mutagenesis, no significant spectroscopic changes on reaction with stoichiometric or higher amounts of H2O2 are seen
Y39A
site-directed mutagenesis, mutation has a destabilizing effect on binding
W191F
-
catalytically inactive mature Ccp1 mutant, Ccp1W191F is a more persistent H2O2 signaling protein than wild-type Ccp1
-
H52L |
-
site-directed mutagenesis, a distal pocket mutant
-
H52Q |
-
site-directed mutagenesis, a distal pocket mutant
-
R48L/W51L/H52L |
-
site-directed mutagenesis, a distal pocket mutant
-
E123D
distal heme pocket mutant, no detectable electrocatalytic turnover of substrate in the highpotential regime
E123Q
distal heme pocket mutant, no detectable electrocatalytic turnover of substrate in the highpotential regime
F102W
distal heme pocket mutant, 10fold decrease in peroxidase activity and high-potential catalytic turnover of hydrogen peroxide
H81G
mutant is highly active
M219Q/F247N
-
site-directed mutagenesis
P75T/H81K/E84Q
-
site-directed mutagenesis
Q113N
distal heme pocket mutant, no detectable electrocatalytic turnover of substrate in the highpotential regime
W191F
-
study of the role of intracomplex dynamics in controlling electron transfer, use of Zn-enzyme in 1:1 complex with cytochrome c
additional information