Comparative Specificities of Two Evolutionarily Divergent Hydrolases Involved in Microbial Degradation of Polychlorinated Biphenyls

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Bibliographische Detailangaben
Zeitschriftentitel:	Journal of Bacteriology
Personen und Körperschaften:	Seah, Stephen Y. K., Labbé, Geneviève, Kaschabek, Stefan R., Reifenrath, Frank, Reineke, Walter, Eltis, Lindsay D.
In:	Journal of Bacteriology, 183, 2001, 5, S. 1511-1516
Format:	E-Article
Sprache:	Englisch
veröffentlicht:	American Society for Microbiology
Schlagwörter:	Molecular Biology Microbiology

Details
Zusammenfassung:	<jats:title>ABSTRACT</jats:title> <jats:p> 2-Hydroxy-6-oxo-6-phenylhexa-2,4-dienoate (HOPDA) hydrolase (BphD) is a key determinant in the aerobic transformation of polychlorinated biphenyls (PCBs) by <jats:italic>Burkholderia</jats:italic> sp. strain LB400 (S. Y. K. Seah, G. Labbé, S. Nerdinger, M. Johnson, V. Snieckus, and L. D. Eltis, J. Biol. Chem. 275:15701–15708, 2000). To determine whether this is also true in divergent biphenyl degraders, the homologous hydrolase of <jats:italic>Rhodococcus globerulus</jats:italic> P6, BphD <jats:sub>P6</jats:sub> , was hyperexpressed, purified to apparent homogeneity, and studied by steady-state kinetics. BphD <jats:sub>P6</jats:sub> hydrolyzed HOPDA with a <jats:italic>k</jats:italic> <jats:sub>cat</jats:sub> / <jats:italic> K <jats:sub>m</jats:sub> </jats:italic> of 1.62 (± 0.03) × 10 <jats:sup>7</jats:sup> M <jats:sup>−1</jats:sup> s <jats:sup>−1</jats:sup> (100 mM phosphate [pH 7.5], 25°C), which is within 70% of that of BphD <jats:sub>LB400</jats:sub> . BphD <jats:sub>P6</jats:sub> was also similar to BphD <jats:sub>LB400</jats:sub> in that it catalyzed the hydrolysis of HOPDAs bearing chloro substituents on the phenyl moiety at least 25 times more specifically than those bearing chloro substituents on the dienoate moiety. However, the rhodococcal enzyme was significantly more specific for 9-Cl and 10-Cl HOPDAs, catalyzing the hydrolysis of 9-Cl, 10-Cl, and 9,10-diCl HOPDAs two- to threefold respectively, more specifically than HOPDA. Moreover, 4-Cl HOPDA competitively inhibited BphD <jats:sub>P6</jats:sub> more effectively than 3-Cl HOPDA, which is the inverse of what was observed in BphD <jats:sub>LB400</jats:sub> . These results demonstrate that BphD is a key determinant in the aerobic transformation of PCBs by divergent biphenyl degraders, but that there exists significant diversity in the specificity of these biphenyl hydrolases. </jats:p>
Umfang:	1511-1516
ISSN:	0021-9193 1098-5530
DOI:	10.1128/jb.183.5.1511-1516.2001