Polychlorinated Biphenyl/Biphenyl Degrading Gene Clusters inRhodococcussp. K37, HA99, and TA431 Are Different from Well-KnownbphGene Clusters of Rhodococci

Abstract

Four kinds of polychlorinated biphenyl (PCB)-degrading Rhodococcus sp. (TA421, TA431, HA99, and K37) have been isolated from termite ecosystem and under alkaline condition. The bph gene cluster involved in the degradation of PCB/biphenyl has been analyzed in strain TA421. This gene cluster was highly homologous to bph gene clusters in R. globerulus P6 and Rhodococcus sp. RHA1. In this study, we cloned and analyzed the bph gene cluster essential to PCB/biphenyl degradation from R. rhodochrous K37. The order of the genes and the sequence were different in K37 than in P6, RHA1, and TA421. The bphC8 K37 gene was more homologous to the meta-cleavage enzyme involved in phenanthrene metabolism than bphC genes involved in biphenyl metabolism. Two other Rhodococcus strains (HA99 and TA431) had PCB/biphenyl degradation gene clusters similar to that in K37. These findings suggest that these bph gene clusters evolved separately from the well-known bph gene clusters of PCB/biphenyl degraders.

Functional Expression of Three Rieske Non-Heme Iron Oxygenases Derived from Actinomycetes inRhodococcusSpecies for Investigation of Their Degradation Capabilities of Dibenzofuran and Chlorinated Dioxins

“…A plasmid, pTB1, that encoded bphA1A2A3A4 of R. erythropolis TA421 was used as the PCR template, 28) and the amplified DNA fragment extending from the start codon of bphA1 TA421 to the stop codon of bphA4 TA421 was ligated downstream of P dfdB (pRK401-P B +bphA TA421 ). The two functionally unidentified genes in the bph gene cluster of R. rhodochrous K37 23) separating the terminal dioxygenase genes bphA1A2 from the electron transfer protein genes bphA3A4 were removed. The DNA fragments for bphA1A2 and bphA3A4 were ligated downstream of the P dfdB , as illustrated in Fig.…”

“…19) However, for some AhDOs derived from high-G+C Gram-positive actinomycetes, it has been difficult to obtain active enzymes in the E. coli protein expression system (our unpublished results and refs. [20][21][22][23].…”

“…YK3 20) and the biphenyl (BP)-utilizing strains Rhodococcus erythropolis TA421 28) and R. rhodochrous K37. 23) In this study, three actinomycetales-derived AhDO genes for which it is difficult to produce functionally active proteins in E. coli were functionally expressed in a Rhodococcus strain using a constitutive expression promoter for the genus Rhodococcus. Their ability to degrade and deplete DF and chlorinated dioxins was analyzed.…”

“…20,21,23,24,26) The genus Rhodococcus has been extensively investigated, especially for its ability to degrade diverse organic compounds. 34) In addition, the presence of a hydrophobic cell envelope 35,36) might provide Rhodococcus with advantages for the degradation of hydrophobic xenobiotics.…”

“…We have reported that both of these genes were involved in BP utilization. 23,28) The amino acid sequences of their respective BphA subunits had only from 35% (BphA4) to 44% (BphA3) identity. Our preliminary attempts to express the two BphAs in E. coli failed (data not shown for BphA TA421 , see ref.…”