Cloning of three 2,3-dihydroxybiphenyl-1,2-dioxygenase genes from Achromobacter sp. BP3 and the analysis of their roles in the biodegradation of biphenyl.

Three 2,3-dihydroxybiphenyl 1,2-dioxygenase genes (designated as bphC1, bphC2 and bphC3) were cloned from a biphenyl-degrading strain Achromobacter sp. BP3. The amino acid sequence of BphC1 and BphC3 had high similarity (>99%) with the reported BphCs, while BphC2 showed relatively low identity (29.51-50.17%) with the reported BphCs, which indicated that bphC2 might be a novel gene. The bphC1, bphC2 and bphC3 genes were expressed in Escherichia coli BL21 and the products were homogenously purified. BphC1, BphC2 and BphC3 displayed maximum activity at 30°C, 30°C and 40°C, respectively. Their optimal catalysis pH was 8.0, 9.0 and 9.0, respectively. BphC1 and BphC2 had higher substrate affinity and catalytic efficiency on 2,3-dihydroxybiphenyl, while BphC3 exhibited these features on aromatic monocyclic substrates. The bphC1 gene was only induced by biphenyl and bphC3 was induced by both biphenyl and toluene, while bphC2 was constitutively expressed in strain BP3. These results suggested that BphC1 and BphC3 played a role in the upstream and downstream metabolic pathways of biphenyl, respectively. However, BphC2 might play a supplementary role and contribute more to the upstream than to the downstream pathway.