Cloning and expression of a novel trans -anethole oxygenase gene from Paraburkholderia sp. MR185

[摘要] trans-Anethole oxygenase (TAO) is the key enzymeresponsible for the oxidation of trans-anethole top-anisaldehyde. A strain, Paraburkholderia sp.MR185, was isolated from soil in Yulin staranise-planting regions using trans-anethole as a solecarbon source and a gene which encodes a proteinwith high similarities to a hypothetical protein ofParaburkholderia sp. MM5384-R2 which shows61.27% identies with TAO from Pseudomonas putidaJYR-1 was cloned and sequenced. The gene, tao,was expressed in E. coli cells and its protein productwas purified by affinity chromatography throughregenerated amorphous cellulose (RAC). SDSPAGE analysis indicated a clear band of recombinant protein TAO, and its molecular weight, 38.3kDa, was consistent with the theoretical value.Its enzyme activity of producing p-anisaldehydefrom trans-anethole was detected by DNPH(2,4-dinitrophenylhydrazine) chromogenic reactionand HPLC, and the specific activity of TAO reached3.93 U/mg protein. Immobilized TAO on RAC wasused to catalyze the production of p-anisaldehydefrom trans-anethole, and the enzyme retained morethan 60% of its initial activity after 10 uses. This isthe first report on Paraburkholderia TAO.

[学科分类] 微生物学和免疫学

[关键词] E. coli;Paraburkholderia;trans-anethole;p-anisaldehyde;trans-anethole oxygenase