Cloning, overexpression, purification, and characterization of the maleylacetate reductase from Sphingobium chlorophenolicum strain ATCC 53874.
Curr Microbiol
; 58(6): 599-603, 2009 Jun.
Article
en En
| MEDLINE
| ID: mdl-19242755
ABSTRACT
The final enzyme in the pentachlorophenol (PCP) biodegradation pathway in Sphingobium chlorophenolicum is maleylacetate reductase (PcpE), which catalyzes the reductive dehalogenation of 2-chloromaleylacetate to maleylacetate and the subsequent reduction of malyelacetate to 3-oxoadipate. In this study, the pcpE gene was cloned from S. chlorophenolicum strain ATCC 53874 and overexpressed in Escherichia coli BL21-AI cells. The recombinant PcpE, purified to higher than 95% purity using affinity chromatography, exhibited optimal activity at pH 7.0. The kinetic parameters k(cat) and K(m) were 1.2 +/- 0.3 s(-1) and 0.09 +/- 0.04 mM, respectively, against maleylacetate under the optimal pH. In addition, the purified PcpE was able to restore PCP-degrading capability to S. chlorophenolicum strain ATCC 39723, implicating that there was no functional PcpE in the ATCC 39723 strain.
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1
Banco de datos:
MEDLINE
Asunto principal:
Proteínas Bacterianas
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Clonación Molecular
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Sphingomonadaceae
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Oxidorreductasas actuantes sobre Donantes de Grupo CH-CH
Idioma:
En
Año:
2009
Tipo del documento:
Article