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Literature summary for 3.5.2.6 extracted from

  • Papp-Wallace, K.M.; Taracila, M.; Hornick, J.M.; Hujer, A.M.; Hujer, K.M.; Distler, A.M.; Endimiani, A.; Bonomo, R.A.
    Substrate selectivity and a novel role in inhibitor discrimination by residue 237 in the KPC-2 beta-lactamase (2010), Antimicrob. Agents Chemother., 54, 2867-2877.
    View publication on PubMedView publication on EuropePMC

Cloned(Commentary)

Cloned (Comment) Organism
expression in Escherichia coli plasmid pBC

Protein Variants

Protein Variants Comment Organism
T237A the mutant enzyme exhibits elevated MICs for ampicillin, piperacillin, and the beta-lactam–beta-lactamase inhibitor combinations. The T237A mutant enzyme demonstrates a lower kcat/Km for imipenem, cephalothin, and cefotaxime. The T237A mutant enzyme shows increased Kis for clavulanic acid, sulbactam, and tazobactam plasmid pBC
T237S the mutant enzyme maintains MICs equivalent to those of the wild type against all of the beta-lactams tested, including carbapenems. The mutant enzyme displays substrate kinetics similar to those of the wild-type KPC-2 enzyme. The T237S mutant enzyme displays Kis similar to wilde-type enzyme plasmid pBC

Inhibitors

Inhibitors Comment Organism Structure
clavulanic acid
-
plasmid pBC
Sulbactam
-
plasmid pBC
tazobactam
-
plasmid pBC

KM Value [mM]

KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
0.005
-
nitrocefin pH 7.4, 22°C, T237S mutant enzyme plasmid pBC
0.006
-
piperacillin pH 7.4, 22°C, T237S mutant enzyme plasmid pBC
0.006
-
nitrocefin pH 7.4, 22°C, wild-type enzyme plasmid pBC
0.012
-
piperacillin pH 7.4, 22°C, T237A mutant enzyme plasmid pBC
0.014
-
cephalothin pH 7.4, 22°C, wild-type enzyme plasmid pBC
0.015
-
nitrocefin pH 7.4, 22°C, T237A mutant enzyme plasmid pBC
0.015
-
piperacillin pH 7.4, 22°C, wild-type enzyme plasmid pBC
0.018
-
cephalothin pH 7.4, 22°C, T237S mutant enzyme plasmid pBC
0.019
-
Imipenem pH 7.4, 22°C, wild-type enzyme plasmid pBC
0.021
-
Imipenem pH 7.4, 22°C, T237S mutant enzyme plasmid pBC
0.03
-
cephalothin pH 7.4, 22°C, T237A mutant enzyme plasmid pBC
0.031
-
Imipenem pH 7.4, 22°C, T237A mutant enzyme plasmid pBC
0.168
-
cefotaxime pH 7.4, 22°C, T237S mutant enzyme plasmid pBC
0.207
-
cefotaxime pH 7.4, 22°C, wild-type enzyme plasmid pBC

Organism

Organism UniProt Comment Textmining
plasmid pBC
-
vector SK(+)-blaKPC-2
-

Purification (Commentary)

Purification (Comment) Organism
-
plasmid pBC

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
cefotaxime + H2O
-
plasmid pBC (2R)-5-[(acetyloxy)methyl]-2-[(R)-[[(2E)-2-(2-amino-1,3-thiazol-4-yl)-2-(methoxyimino)acetyl]amino](carboxy)methyl]-3,6-dihydro-2H-1,3-thiazine-4-carboxylic acid
-
?
cephalothin + H2O
-
plasmid pBC (2R)-5-[(acetyloxy)methyl]-2-[(R)-carboxy[(thiophen-2-ylacetyl)amino]methyl]-3,6-dihydro-2H-1,3-thiazine-4-carboxylic acid
-
?
imipenem + H2O
-
plasmid pBC (5R)-3-[[2-(carbonoimidoylamino)ethyl]sulfanyl]-5-[(1S,2R)-1-carboxy-2-hydroxypropyl]-4,5-dihydro-1H-pyrrole-2-carboxylic acid
-
?
nitrocefin + H2O
-
plasmid pBC (2R)-2-[(R)-carboxy[(thiophen-2-ylacetyl)amino]methyl]-5-[(E)-2-(2,4-dinitrophenyl)ethenyl]-3,6-dihydro-2H-1,3-thiazine-4-carboxylic acid
-
?
piperacillin + H2O
-
plasmid pBC (2R,4S)-2-[(R)-carboxy[[(2R)-2-[[(4-ethyl-2,3-dioxopiperazin-1-yl)carbonyl]amino]-2-phenylacetyl]amino]methyl]-5,5-dimethyl-1,3-thiazolidine-4-carboxylic acid
-
?

Synonyms

Synonyms Comment Organism
KPC-2 beta-lactamase
-
plasmid pBC

Turnover Number [1/s]

Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
6.8
-
Imipenem pH 7.4, 22°C, T237A mutant enzyme plasmid pBC
19
-
Imipenem pH 7.4, 22°C, wild-type enzyme plasmid pBC
21
-
Imipenem pH 7.4, 22°C, T237S mutant enzyme plasmid pBC
26
-
cephalothin pH 7.4, 22°C, T237A mutant enzyme plasmid pBC
42
-
piperacillin pH 7.4, 22°C, T237A mutant enzyme plasmid pBC
42
-
piperacillin pH 7.4, 22°C, T237S mutant enzyme plasmid pBC
43
-
cefotaxime pH 7.4, 22°C, T237S mutant enzyme plasmid pBC
44
-
cephalothin pH 7.4, 22°C, wild-type enzyme plasmid pBC
50
-
nitrocefin pH 7.4, 22°C, T237S mutant enzyme plasmid pBC
51
-
piperacillin pH 7.4, 22°C, wild-type enzyme plasmid pBC
52
-
nitrocefin pH 7.4, 22°C, wild-type enzyme plasmid pBC
65
-
cephalothin pH 7.4, 22°C, T237S mutant enzyme plasmid pBC
142
-
cefotaxime pH 7.4, 22°C, wild-type enzyme plasmid pBC
191
-
nitrocefin pH 7.4, 22°C, T237A mutant enzyme plasmid pBC

Ki Value [mM]

Ki Value [mM] Ki Value maximum [mM] Inhibitor Comment Organism Structure
0.011
-
clavulanic acid pH 7.4, 22°C, wild-type enzyme plasmid pBC
0.013
-
clavulanic acid pH 7.4, 22°C, T237S mutant enzyme plasmid pBC
0.053
-
tazobactam pH 7.4, 22°C, T237S mutant enzyme plasmid pBC
0.055
-
clavulanic acid pH 7.4, 22°C, T237A mutant enzyme plasmid pBC
0.074
-
tazobactam pH 7.4, 22°C, wild-type enzyme plasmid pBC
0.097
-
Sulbactam pH 7.4, 22°C, T237S mutant enzyme plasmid pBC
0.167
-
Sulbactam pH 7.4, 22°C, wild-type enzyme plasmid pBC
0.174
-
tazobactam pH 7.4, 22°C, T237A mutant enzyme plasmid pBC
0.703
-
Sulbactam pH 7.4, 22°C, T237A mutant enzyme plasmid pBC

kcat/KM [mM/s]

kcat/KM Value [1/mMs-1] kcat/KM Value Maximum [1/mMs-1] Substrate Comment Organism Structure
220
-
Imipenem pH 7.4, 22°C, T237A mutant enzyme plasmid pBC
260
-
cefotaxime pH 7.4, 22°C, T237S mutant enzyme plasmid pBC
700
-
cefotaxime pH 7.4, 22°C, wild-type enzyme plasmid pBC
900
-
cephalothin pH 7.4, 22°C, T237A mutant enzyme plasmid pBC
1000
-
Imipenem pH 7.4, 22°C, T237S mutant enzyme plasmid pBC
1000
-
Imipenem pH 7.4, 22°C, wild-type enzyme plasmid pBC
3100
-
cephalothin pH 7.4, 22°C, wild-type enzyme plasmid pBC
3400
-
piperacillin pH 7.4, 22°C, wild-type enzyme plasmid pBC
3500
-
piperacillin pH 7.4, 22°C, T237A mutant enzyme plasmid pBC
3600
-
cephalothin pH 7.4, 22°C, T237S mutant enzyme plasmid pBC
4000
-
piperacillin pH 7.4, 22°C, T237S mutant enzyme plasmid pBC
8700
-
nitrocefin pH 7.4, 22°C, wild-type enzyme plasmid pBC
10000
-
nitrocefin pH 7.4, 22°C, T237S mutant enzyme plasmid pBC
13000
-
nitrocefin pH 7.4, 22°C, T237A mutant enzyme plasmid pBC