亚洲传统医药

Abstract: In our previous work, new wild-type strain (WT16) that exhibited nitrilase activity toward 3-cyanopyridine was isolated from soil through enrichment culture using acetonitrile or 3-cyanopyridine as the sole source of nitrogen. To improve the strain’s nitrilase activity for industrial application, it was further subjected to mutagenesis using LiCl, and the mutant strain Agrobacterium sp. SPUA14 was obtained. The nitrilase activity of Agrobacterium sp. SPUA14 was markedly increased by 50.6% compared with WT16 (26.23 μmol·g-1·h-1). The nitrilase from Agrobacterium sp.SPUA14 was subsequently amplified and expressed in E. coli BL21 (DE3). The apparent Km and rmax of the nitrilase against 3-cyanopyridine were 8.35×10-2 mol·L-1 and 3.575× 10-3 mol·L-1·min-1, respectively. After 40 min reaction using E. coli BL21/pET22b-a14 resting cells, 3-cyanopyridine at the concentration of 10 mmol·L-1 was completely hydrolyzed and 91.2% conversion rates were attained. The determination of the substrate tolerance indicated that the optimum substrate concentration of nitrilase expressed in the engineered bacteria is approximately 300 mmol·L-1, which makes it a potential biocatalyst for the production of nicotinic acid.

Key words: nitrilase, 3-cyanopyridine, Agrobacterium sp, biotransformation