STUDIES ON THE MOLECULAR CLONING OF A GENE FOR ARYLETHERASE FROM A LIGNINOLYTIC ERWINIA SP. (LIGNIN, BIODEGRADATION, BACTERIAL).

摘要

Recombinant DNA techniques were utilized to clone a gene from the ligninolytic system of a saprophytic Erwinia sp. into Escherichia coli using pBR322 as a vector. The cloned gene encodes aryletherase activity and was selected from amp('r) tet('S) transformants of E. coli Cs412 using salicin as the selective carbon source. This compound has structural characteristics similar to some lignin model compounds. The cloned gene enclodes activity which oxidizes aromatic lignin model compounds resulting ultimately in the breakage of arylether bonds while leaving the aromatic nucleus intact. At least one lower molecular weight aromatic compound is also released from kraft lignin upon incubation with whole cells harboring the recombinant plasmid, pNC1. Restriction enzyme analysis revealed that pNC1 contains an insert of approximately 810 bp of Erwinia sp. DNA. Southern blot hybridizations indicate that the gene is present as a single copy on the Erwinia sp. genome. Dideoxynucleotide DNA sequencing suggests that the gene is transcribed by readthrough of transcription initiated at the tetracycline resistance promoter in pNC1. The predicted translation product has an estimated molecular weight of 21,000 daltons.