SEQUENCE OF THE BPHD GENE ENCODING 2-HYDROXY-6-OXO-(PHENYL CHLOROPHENYL)HEXA-2,4-DIENOIC ACID (HOP/CPDA) HYDROLASE INVOLVED IN THE BIPHENL/POLYCHLORINATED BIPHENYL DEGRADATION PATHWAY IN COMMAMONAS-TESTOSTERONI - EVIDENCE SUGGESTING INVOLVEMENT OF SER(112) IN CATALYTIC ACTIVITY/

The nucleotide sequence of bphD, encoding 2-hydroxy-6-oxo-(phenyl/chlo rophenyl) hexa-2,4-dienoic acid hydrolase involved in the biphenyl/pol ychlorinated biphenyl degradation pathway of Comamonas testosteroni st rain B-356, was determined. Comparison of the deduced amino-acid seque nce with published sequences led to the identification of a 'lipase bo x', containing a consensus pentapeptide sequence GlyXaaSerXaaGly. This suggested that the mechanism of action of this enzyme may involve an Asp-Ser-His catalytic triad similar to that of classical lipases and s erine hydrolases. Further biochemical and genetic evidence for the act ive-site involvement of Ser(112) was obtained by showing that a semipu rified enzyme was inhibited by PMSF, a classic inhibitor of serine hyd rolases, and by site-directed Ser(112)-->Ala mutagenesis.