Literature summary for 3.5.2.6 extracted from

Bebrone, C.; Anne, C.; Kerff, F.; Garau, G.; De Vriendt, K.; Lantin, R.; Devreese, B.; Van Beeumen, J.; Dideberg, O.; Frere, J.; Galleni, M.
Mutational analysis of the zinc- and substrate-binding sites in the CphA metallo-beta -lactamase from Aeromonas hydrophila (2008), Biochem. J., 414, 151-159.

Cloned(Commentary)

Cloned (Comment)	Organism
expressed in Escherichia coli	Aeromonas hydrophila

Protein Variants

Protein Variants	Comment	Organism
D120A	site-directed mutant of CphA, generated and characterized	Aeromonas hydrophila
D120T	site-directed mutant of CphA, generated and characterized	Aeromonas hydrophila
H118A	site-directed mutant of CphA, generated and characterized, H118A and H196A mutations lead to a decrease of at least 1000fold in activity against carbapenems compared with the wild-type enzyme	Aeromonas hydrophila
H196A	site-directed mutant of CphA, generated and characterized, H118A and H196A mutations lead to a decrease of at least 1000fold in activity against carbapenems compared with the wild-type enzyme	Aeromonas hydrophila
H263A	site-directed mutant of CphA, generated and characterized	Aeromonas hydrophila
K224Q	site-directed mutant of CphA, generated and characterized	Aeromonas hydrophila
K226Q	site-directed mutant of CphA, generated and characterized	Aeromonas hydrophila
N116H/N220G	site-directed mutant of CphA, generated and characterized	Aeromonas hydrophila
N116H/N220G/K224Q	site-directed mutant of CphA, generated and characterized	Aeromonas hydrophila
T157A	site-directed mutant of CphA, generated and characterized	Aeromonas hydrophila
V67A	site-directed mutant of CphA, generated and characterized	Aeromonas hydrophila
V67D	site-directed mutant of CphA, generated and characterized	Aeromonas hydrophila
V67I	site-directed mutant of CphA, generated and characterized	Aeromonas hydrophila
V67LFKHV	site-directed mutant of CphA, generated and characterized	Aeromonas hydrophila

Inhibitors

Inhibitors	Comment	Organism	Structure
EDTA	inactivation of CphA wild-type and the H118A, H196A, H263A and D120A mutants by EDTA is studied in 15 mM sodium cacodylate (pH 6.5) at 30°C in the presence of different concentrations of EDTA	Aeromonas hydrophila

KM Value [mM]

KM Value [mM]	KM Value Maximum [mM]	Substrate	Comment	Organism
0.004	-	nitrocefin	N116H/N220G mutant, 30°C, pH 6.5	Aeromonas hydrophila
0.015	-	nitrocefin	H196A mutant, 30°C, pH 6.5	Aeromonas hydrophila
0.015	-	nitrocefin	H263A mutant, 30°C, pH 6.5	Aeromonas hydrophila
0.026	-	nitrocefin	D120A mutant, 30°C, pH 6.5	Aeromonas hydrophila
0.026	-	nitrocefin	V67A mutant, 30°C, pH 6.5	Aeromonas hydrophila
0.03	-	nitrocefin	H118A mutant, 30°C, pH 6.5	Aeromonas hydrophila
0.032	-	nitrocefin	N116H/N220G/K224Q mutant, 30°C, pH 6.5	Aeromonas hydrophila
0.036	-	nitrocefin	V67D mutant, 30°C, pH 6.5	Aeromonas hydrophila
0.05	-	nitrocefin	D120T mutant, 30°C, pH 6.5	Aeromonas hydrophila
0.05	-	cefotaxime	N116H/N220G mutant, 30°C, pH 6.5	Aeromonas hydrophila
0.05	-	nitrocefin	V67I mutant, 30°C, pH 6.5	Aeromonas hydrophila
0.053	-	cephaloridine	N116H/N220G/K224Q mutant, 30°C, pH 6.5	Aeromonas hydrophila
0.1	-	cefotaxime	above, wild-type, 30°C, pH 6.5	Aeromonas hydrophila
0.1	-	biapenem	D120A mutant, 30°C, pH 6.5	Aeromonas hydrophila
0.1	-	biapenem	N116H/N220G mutant, 30°C, pH 6.5	Aeromonas hydrophila
0.1	-	biapenem	V67A mutant, 30°C, pH 6.5	Aeromonas hydrophila
0.11	-	nitrocefin	T157A mutant, 30°C, pH 6.5	Aeromonas hydrophila
0.135	-	nitrocefin	K224Q mutant, 30°C, pH 6.5	Aeromonas hydrophila
0.14	-	cefotaxime	K226Q mutant, 30°C, pH 6.5	Aeromonas hydrophila
0.145	-	cephaloridine	N116H/N220G mutant, 30°C, pH 6.5	Aeromonas hydrophila
0.15	-	benzylpenicillin	N116H/N220G mutant, 30°C, pH 6.5	Aeromonas hydrophila
0.17	-	biapenem	wild-type, 30°C, pH 6.5	Aeromonas hydrophila
0.18	-	Imipenem	V67I mutant, 30°C, pH 6.5	Aeromonas hydrophila
0.185	-	Imipenem	N116H/N220G mutant, 30°C, pH 6.5	Aeromonas hydrophila
0.21	-	Imipenem	D120T mutant, 30°C, pH 6.5	Aeromonas hydrophila
0.21	-	benzylpenicillin	H118A mutant, 30°C, pH 6.5	Aeromonas hydrophila
0.22	-	Imipenem	V67A mutant, 30°C, pH 6.5	Aeromonas hydrophila
0.3	-	Imipenem	N116H/N220G/K224Q mutant, 30°C, pH 6.5	Aeromonas hydrophila
0.31	-	Imipenem	V67D mutant, 30°C, pH 6.5	Aeromonas hydrophila
0.33	-	Imipenem	D120A mutant, 30°C, pH 6.5	Aeromonas hydrophila
0.34	-	Imipenem	wild-type, 30°C, pH 6.5	Aeromonas hydrophila
0.38	-	Imipenem	K226Q mutant, 30°C, pH 6.5	Aeromonas hydrophila
0.39	-	biapenem	N116H/N220G/K224Q mutant, 30°C, pH 6.5	Aeromonas hydrophila
0.434	-	cefotaxime	N116H/N220G/K224Q mutant, 30°C, pH 6.5	Aeromonas hydrophila
0.45	-	cefotaxime	T157A mutant, 30°C, pH 6.5	Aeromonas hydrophila
0.55	-	benzylpenicillin	T157A mutant, 30°C, pH 6.5	Aeromonas hydrophila
0.57	-	Imipenem	T157A mutant, 30°C, pH 6.5	Aeromonas hydrophila
0.7	-	cefotaxime	K224Q mutant, 30°C, pH 6.5	Aeromonas hydrophila
0.73	-	biapenem	H118A mutant, 30°C, pH 6.5	Aeromonas hydrophila
0.73	-	benzylpenicillin	V67I mutant, 30°C, pH 6.5	Aeromonas hydrophila
0.8	-	benzylpenicillin	K226Q mutant, 30°C, pH 6.5	Aeromonas hydrophila
0.84	-	biapenem	K226Q mutant, 30°C, pH 6.5	Aeromonas hydrophila
0.85	-	nitrocefin	K226Q mutant, 30°C, pH 6.5	Aeromonas hydrophila
0.87	-	benzylpenicillin	wild-type, 30°C, pH 6.5	Aeromonas hydrophila
0.92	-	benzylpenicillin	N116H/N220G/K224Q mutant, 30°C, pH 6.5	Aeromonas hydrophila
1	-	cephaloridine	K224Q mutant, 30°C, pH 6.5	Aeromonas hydrophila
1	-	benzylpenicillin	V67A mutant, 30°C, pH 6.5	Aeromonas hydrophila
1.15	-	Imipenem	H118A mutant, 30°C, pH 6.5	Aeromonas hydrophila
1.3	-	nitrocefin	wild-type, 30°C, pH 6.5	Aeromonas hydrophila
1.5	-	biapenem	above, H263A mutant, 30°C, pH 6.5	Aeromonas hydrophila
1.5	-	Imipenem	above, K224Q mutant, 30°C, pH 6.5	Aeromonas hydrophila
1.5	-	biapenem	above, K224Q mutant, 30°C, pH 6.5	Aeromonas hydrophila
1.5	-	biapenem	above, T157A mutant, 30°C, pH 6.5	Aeromonas hydrophila
1.5	-	biapenem	above, V67D mutant, 30°C, pH 6.5	Aeromonas hydrophila
1.5	-	biapenem	above, V67I mutant, 30°C, pH 6.5	Aeromonas hydrophila
1.5	-	Imipenem	above, V67LFKHV mutant, 30°C, pH 6.5	Aeromonas hydrophila
1.5	-	biapenem	above, V67LFKHV mutant, 30°C, pH 6.5	Aeromonas hydrophila
1.7	-	Imipenem	H263A mutant, 30°C, pH 6.5	Aeromonas hydrophila
1.7	-	cephaloridine	K226Q mutant, 30°C, pH 6.5	Aeromonas hydrophila
2	-	nitrocefin	above, V67LFKHV mutant, 30°C, pH 6.5	Aeromonas hydrophila
4.3	-	Imipenem	H196A mutant, 30°C, pH 6.5	Aeromonas hydrophila
4.5	-	cephaloridine	above, H118A mutant, 30°C, pH 6.5	Aeromonas hydrophila
4.5	-	cephaloridine	above, H196A mutant, 30°C, pH 6.5	Aeromonas hydrophila
5	-	benzylpenicillin	above, H196A mutant, 30°C, pH 6.5	Aeromonas hydrophila
5	-	benzylpenicillin	above, K224Q mutant, 30°C, pH 6.5	Aeromonas hydrophila
5	-	benzylpenicillin	above, V67D mutant, 30°C, pH 6.5	Aeromonas hydrophila
6	-	cephaloridine	wild-type, 30°C, pH 6.5	Aeromonas hydrophila

Metals/Ions

Metals/Ions	Comment	Organism	Structure
Zn2+	zinc content is determined by ICP-MS. Position of the second inhibitory zinc ion has not been identified	Aeromonas hydrophila

Molecular Weight [Da]

Molecular Weight [Da]	Molecular Weight Maximum [Da]	Comment	Organism
25120	-	H118A mutant, denatured, calculated mass	Aeromonas hydrophila
25120	-	H196A mutant, denatured, calculated mass	Aeromonas hydrophila
25120	-	H263A mutant, denatured, calculated mass	Aeromonas hydrophila
25120	-	H263A mutant, denatured, measured mass	Aeromonas hydrophila
25130	-	H118A mutant, denatured, measured mass	Aeromonas hydrophila
25130	-	H196A mutant, denatured, measured mass	Aeromonas hydrophila
25150	-	D120A mutant, denatured, calculated mass	Aeromonas hydrophila
25150	-	D120A mutant, denatured, measured mass	Aeromonas hydrophila
25160	-	N116H/R121H/K224Q mutant, denatured, calculated mass	Aeromonas hydrophila
25160	-	N116H/R121H/K224Q mutant, denatured, measured mass	Aeromonas hydrophila
25160	-	T157A mutant, denatured, calculated mass	Aeromonas hydrophila
25160	-	T157A mutant, denatured, measured mass	Aeromonas hydrophila
25160	-	V67A mutant, denatured, calculated mass	Aeromonas hydrophila
25160	-	V67A mutant, denatured, measured mass	Aeromonas hydrophila
25180	-	D120T mutant, denatured, calculated mass	Aeromonas hydrophila
25180	-	D120T mutant, denatured, measured mass	Aeromonas hydrophila
25190	-	H118A mutant, native, measured mass	Aeromonas hydrophila
25190	-	H196A mutant, native, measured mass	Aeromonas hydrophila
25190	-	K224Q mutant, denatured, calculated mass	Aeromonas hydrophila
25190	-	K224Q mutant, denatured, measured mass	Aeromonas hydrophila
25190	-	K226Q mutant, denatured, calculated mass	Aeromonas hydrophila
25190	-	K226Q mutant, denatured, measured mass	Aeromonas hydrophila
25190	-	wild-type, denatured, calculated mass	Aeromonas hydrophila
25190	-	wild-type, denatured, measured mass	Aeromonas hydrophila
25200	-	V67I mutant, denatured, calculated mass	Aeromonas hydrophila
25200	-	V67I mutant, denatured, measured mass	Aeromonas hydrophila
25210	-	D120A mutant, native, measured mass	Aeromonas hydrophila
25220	-	H263A mutant, native, measured mass	Aeromonas hydrophila
25220	-	N116H/R121H/K224Q mutant, native, measured mass	Aeromonas hydrophila
25220	-	T157A mutant, native, measured mass	Aeromonas hydrophila
25220	-	V67A mutant, native, measured mass	Aeromonas hydrophila
25240	-	D120T mutant, native, measured mass	Aeromonas hydrophila
25250	-	K224Q mutant, native, measured mass	Aeromonas hydrophila
25250	-	K226Q mutant, native, measured mass	Aeromonas hydrophila
25250	-	wild-type, native, measured mass	Aeromonas hydrophila
25270	-	V67I mutant, native, measured mass	Aeromonas hydrophila
25810	-	V67LFKHV mutant, denatured, measured mass	Aeromonas hydrophila
25880	-	V67LFKHV mutant, native, measured mass, 1 Zn2+-ion	Aeromonas hydrophila
25900	-	V67LFKHV mutant, denatured, calculated mass	Aeromonas hydrophila
25940	-	V67LFKHV mutant, native, measured mass, 2 Zn2+-ions	Aeromonas hydrophila

Organism

Organism	UniProt	Comment	Textmining
Aeromonas hydrophila	P26918	-	-

Purification (Commentary)

Purification (Comment)	Organism
-	Aeromonas hydrophila

Substrates and Products (Substrate)

Substrates	Comment Substrates	Organism	Products	Comment (Products)	Rev.
benzylpenicillin + H2O	-	Aeromonas hydrophila	(2R,4S)-2-[(R)-carboxy(2-phenylacetamido)methyl]-5,5-dimethyl-1,3-thiazolidine-4-carboxylic acid	-	?
biapenem + H2O	-	Aeromonas hydrophila	(4R,5S)-5-[(1S,2R)-1-carboxy-2-hydroxypropyl]-3-[(6,7-dihydro-5H-pyrazolo[1,2-a][1,2,4]triazol-4-ium-6-yl)sulfanyl]-4-methyl-4,5-dihydro-1H-pyrrole-2-carboxylate	-	?
carbapenem + H2O	-	Aeromonas hydrophila	[(2R)-2,3-dihydro-1H-pyrrol-2-yl]acetic acid	-	?
cefotaxime + H2O	-	Aeromonas hydrophila	(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	-	?
cephaloridine + H2O	-	Aeromonas hydrophila	(2R)-2-[(R)-carboxy[(thiophen-2-ylacetyl)amino]methyl]-5-(pyridinium-1-ylmethyl)-3,6-dihydro-2H-1,3-thiazine-4-carboxylate	-	?
imipenem + H2O	-	Aeromonas hydrophila	(5R)-5-[(1S,2R)-1-carboxy-2-hydroxypropyl]-3-({2-[(iminomethyl)amino]ethyl}sulfanyl)-4,5-dihydro-1H-pyrrole-2-carboxylic acid	-	?
nitrocefin + H2O	-	Aeromonas hydrophila	(2R)-2-{(R)-carboxy[2-(thiophen-2-yl)acetamido]methyl}-5-[(E)-2-(2,4-dinitrophenyl)ethenyl]-3,6-dihydro-2H-1,3-thiazine-4-carboxylic acid	-	?

Subunits

Subunits	Comment	Organism
More	His118 and His196 participate in the catalytic mechanism of the subclass B2 CphA metallo-beta-lactamase, for which an intact Asp120/Cys221/His263 site is essential	Aeromonas hydrophila

Synonyms

Synonyms	Comment	Organism
carbapenemase	-	Aeromonas hydrophila
CphA	-	Aeromonas hydrophila
metallo-beta-lactamase	-	Aeromonas hydrophila

Turnover Number [1/s]

Turnover Number Minimum [1/s]	Turnover Number Maximum [1/s]	Substrate	Comment	Organism
0.0002	-	cefotaxime	above, wild-type, 30°C, pH 6.5	Aeromonas hydrophila
0.0002	-	nitrocefin	D120T mutant, 30°C, pH 6.5	Aeromonas hydrophila
0.0006	-	cefotaxime	K226Q mutant, 30°C, pH 6.5	Aeromonas hydrophila
0.001	-	cefotaxime	K224Q mutant, 30°C, pH 6.5	Aeromonas hydrophila
0.0015	-	biapenem	D120A mutant, 30°C, pH 6.5	Aeromonas hydrophila
0.0024	-	nitrocefin	D120A mutant, 30°C, pH 6.5	Aeromonas hydrophila
0.003	-	nitrocefin	H263A mutant, 30°C, pH 6.5	Aeromonas hydrophila
0.004	-	benzylpenicillin	K226Q mutant, 30°C, pH 6.5	Aeromonas hydrophila
0.004	-	benzylpenicillin	V67A mutant, 30°C, pH 6.5	Aeromonas hydrophila
0.004	-	nitrocefin	V67D mutant, 30°C, pH 6.5	Aeromonas hydrophila
0.004	-	nitrocefin	V67I mutant, 30°C, pH 6.5	Aeromonas hydrophila
0.005	-	biapenem	above, H263A mutant, 30°C, pH 6.5	Aeromonas hydrophila
0.005	-	nitrocefin	H118A mutant, 30°C, pH 6.5	Aeromonas hydrophila
0.005	-	nitrocefin	V67A mutant, 30°C, pH 6.5	Aeromonas hydrophila
0.006	-	nitrocefin	H196A mutant, 30°C, pH 6.5	Aeromonas hydrophila
0.006	-	nitrocefin	above, V67LFKHV mutant, 30°C, pH 6.5	Aeromonas hydrophila
0.006	-	cephaloridine	below, wild-type, 30°C, pH 6.5	Aeromonas hydrophila
0.006	-	cephaloridine	K226Q mutant, 30°C, pH 6.5	Aeromonas hydrophila
0.007	-	benzylpenicillin	H118A mutant, 30°C, pH 6.5	Aeromonas hydrophila
0.007	-	nitrocefin	K224Q mutant, 30°C, pH 6.5	Aeromonas hydrophila
0.008	-	nitrocefin	wild-type, 30°C, pH 6.5	Aeromonas hydrophila
0.01	-	cephaloridine	above, H118A mutant, 30°C, pH 6.5	Aeromonas hydrophila
0.01	-	nitrocefin	K226Q mutant, 30°C, pH 6.5	Aeromonas hydrophila
0.015	-	cefotaxime	N116H/N220G/K224Q mutant, 30°C, pH 6.5	Aeromonas hydrophila
0.015	-	nitrocefin	T157A mutant, 30°C, pH 6.5	Aeromonas hydrophila
0.02	-	Imipenem	H263A mutant, 30°C, pH 6.5	Aeromonas hydrophila
0.023	-	cefotaxime	T157A mutant, 30°C, pH 6.5	Aeromonas hydrophila
0.024	-	Imipenem	D120A mutant, 30°C, pH 6.5	Aeromonas hydrophila
0.024	-	cephaloridine	K224Q mutant, 30°C, pH 6.5	Aeromonas hydrophila
0.025	-	Imipenem	D120T mutant, 30°C, pH 6.5	Aeromonas hydrophila
0.03	-	benzylpenicillin	wild-type, 30°C, pH 6.5	Aeromonas hydrophila
0.03	-	benzylpenicillin	above, K224Q mutant, 30°C, pH 6.5	Aeromonas hydrophila
0.03	-	benzylpenicillin	above, V67D mutant, 30°C, pH 6.5	Aeromonas hydrophila
0.03	-	cephaloridine	N116H/N220G/K224Q mutant, 30°C, pH 6.5	Aeromonas hydrophila
0.08	-	benzylpenicillin	V67I mutant, 30°C, pH 6.5	Aeromonas hydrophila
0.1	-	benzylpenicillin	above, H196A mutant, 30°C, pH 6.5	Aeromonas hydrophila
0.1	-	biapenem	H118A mutant, 30°C, pH 6.5	Aeromonas hydrophila
0.11	-	cephaloridine	above, H196A mutant, 30°C, pH 6.5	Aeromonas hydrophila
0.13	-	benzylpenicillin	N116H/N220G/K224Q mutant, 30°C, pH 6.5	Aeromonas hydrophila
0.2	-	nitrocefin	N116H/N220G/K224Q mutant, 30°C, pH 6.5	Aeromonas hydrophila
0.26	-	biapenem	N116H/N220G/K224Q mutant, 30°C, pH 6.5	Aeromonas hydrophila
0.3	-	cefotaxime	N116H/N220G mutant, 30°C, pH 6.5	Aeromonas hydrophila
0.7	-	nitrocefin	N116H/N220G mutant, 30°C, pH 6.5	Aeromonas hydrophila
0.9	-	cephaloridine	N116H/N220G mutant, 30°C, pH 6.5	Aeromonas hydrophila
0.9	-	Imipenem	N116H/N220G/K224Q mutant, 30°C, pH 6.5	Aeromonas hydrophila
1	-	benzylpenicillin	T157A mutant, 30°C, pH 6.5	Aeromonas hydrophila
2.4	-	biapenem	above, V67LFKHV mutant, 30°C, pH 6.5	Aeromonas hydrophila
2.4	-	Imipenem	H118A mutant, 30°C, pH 6.5	Aeromonas hydrophila
3.3	-	benzylpenicillin	N116H/N220G mutant, 30°C, pH 6.5	Aeromonas hydrophila
5	-	biapenem	N116H/N220G mutant, 30°C, pH 6.5	Aeromonas hydrophila
16	-	Imipenem	H196A mutant, 30°C, pH 6.5	Aeromonas hydrophila
16	-	Imipenem	N116H/N220G mutant, 30°C, pH 6.5	Aeromonas hydrophila
30	-	Imipenem	above, V67LFKHV mutant, 30°C, pH 6.5	Aeromonas hydrophila
50	-	biapenem	above, V67I mutant, 30°C, pH 6.5	Aeromonas hydrophila
60	-	biapenem	above, K224Q mutant, 30°C, pH 6.5	Aeromonas hydrophila
100	-	biapenem	above, V67D mutant, 30°C, pH 6.5	Aeromonas hydrophila
100	-	biapenem	V67A mutant, 30°C, pH 6.5	Aeromonas hydrophila
130	-	biapenem	above, T157A mutant, 30°C, pH 6.5	Aeromonas hydrophila
300	-	biapenem	wild-type, 30°C, pH 6.5	Aeromonas hydrophila
490	-	Imipenem	above, K224Q mutant, 30°C, pH 6.5	Aeromonas hydrophila
1000	-	Imipenem	K226Q mutant, 30°C, pH 6.5	Aeromonas hydrophila
1000	-	biapenem	K226Q mutant, 30°C, pH 6.5	Aeromonas hydrophila
1000	-	Imipenem	V67A mutant, 30°C, pH 6.5	Aeromonas hydrophila
1200	-	Imipenem	wild-type, 30°C, pH 6.5	Aeromonas hydrophila
1280	-	Imipenem	V67D mutant, 30°C, pH 6.5	Aeromonas hydrophila
1680	-	Imipenem	T157A mutant, 30°C, pH 6.5	Aeromonas hydrophila
1770	-	Imipenem	V67I mutant, 30°C, pH 6.5	Aeromonas hydrophila