4-Chlorobiphenyl을 분해하는 Pseudomonas sp. P20의 pcb 유전자군의 클로닝 = Cloning of pcb Genes in Pseudomonas sp. P20 Specifying Degradation of 4-Chlorobiphenyl

국문 초록 (Abstract)

Pseudomonas sp. P2O은 난분해성 방향족 탄화수소중의 한가지인 4-chlorobiphenyl(4CB)을 meta-cleavage를 거쳐 4-chlorobenzoic acid로 분해하는 세균이다. 이 세균의 chromosomal DNA로부터 14 kb의 EcoRI 절편을 pBluescript SK(+) vector를 이용하여 E. coli XL1-Blue에 클로닝하여 4CB 분해유전자를 포함하는 재조합 plasmid인 pCK1과 재조합균주인 E. coli CK1을 얻었다. E. coli CK1은 pcbAB 산물에 의하여 4CB를 분해하고 또 pcbCD 산물에 의하여 2,3-dihydroxybiphenyl(2,3-DHBP)을 분해하여 meta-clevage compound(MCP)와 benzoate를 생성하는 것을 resting cell assay에 의하여 확인하였다. pcbC의 발현은 Pseudomonas sp. P20과 재조합균주인 E. coli CK1에서 모두 지수말기에 가장 잘 되었으며, E. coli CK1에서 pcbC 유전자의 산물인 2,3-DHBP dioxygenase 활성은 Pseudomonas sp. P20에 비하여 2배 이상 높았다. 또 2,3-DHBP dioxygenase는 catechol과 3-methylcatechol을 기질로 했을 때 2,3-DHBP에 비하여 26∼31%의 활성을 나타내었으며, 4-methylcatechol과 4-chlorocatechol에 대해서는 활성이 없었다.

다국어 초록 (Multilingual Abstract)

Pseudomonas sp. P2O was a bacterial isolate which has the ability to degrade 4-chlorobiphenyl(4CB) to 4-chlorobenzoic acid via the process of meta-cleavage. The recombinant plasmid pCK1 was constructed by inserting the 14-kb EcoRI fragment of the chromosomal DNA containing the 4CB-degrading genes into the vector pBluescript SK(+). Subsequently, E. coli XL1-Blue was transformed with the hybrid plasmid producing the recombinant E. coli CK1. The recombinant cells degraded 4CB and 2,3-dihydroxybiphenyl(2,3-DHBP) by the pcbAB and pcbCD gene products, respectively. The pcbC gene was expressed most abundantly at the late exponential phase in E. coli CK1 as well as in Pseudomonas sp. P2O, and the level of the pcbC gene product, 2,3-DHBP dioxygenase, expressed in E. coli CK1 was about two-times higher than in Pseudomonas sp. P2O. The activities of 2,3-DHBP dioxygenase on catechol and 3-methylcatechol were about 26 to 31% of its activity on 2,3-DHBP, but the enzyme did not reveal any activities on 4-methylcatechol and 4-chlorocatechol.