Characterization of Two meta-Cleavage Enzymes from Thermophilic PCB-Degrader Bacillus sp. JF8
Hiroki HIRABAYASHIa, Umechiyo TOKUMOTOa, Takashi HATTA*a, Hohzoh KIYOHARAa, Minoru SHIMURAb, and Kazuhide KIMBARAb
a Research Institute of Technology, Okayama University of Science, 401-1 Seki, 0kayama 703, Japan
b Environmental Biotechnology Laboratory, Railway Technical Research Institute, 2-8-38 Hikaricho, Kokubunji, Tokyo, 185, Japan
Thermophilic Bacillus sp. JF8 degrades up to trichlorinated biphenyls (PCBs) at 60℃. We cloned the genes encoding two types of meta-cleavage enzymes from Bacillus sp. JF8. In this study, we report the sequence analysis and enzymatic properties of these enzymes.
From the gene bank of total DNA of strain JF8, two positive clones, which cleave 2,3-dihydroxybiphenyl (23DHBP) to 2-hydroxy76-oxo-6-phenylhexa-2,4- dienoic acid, were obtained. From substrate specificity studies, one was identified as a bphC gene which was more active for 23DHBP and the other was xylE gene which was active for catechol. Amino acid sequences of BphC and XylE had low homology to other known enzymes from mesophilic PCB-degrading bacteria.
BphC of JF8 contained weak relative activities toward catechol, 3- methylcatechol, 4-methylcatechol, and 4-chlorocatechol compared with activity toward 23DHBP. On the other hand, XylE of JF8 contained significantly low activity toward 3-methylcatechol compared with activity toward catechol.
Optimal pH of BphC and XylE was 8.5 and 7.5, respectively.
