关键词: 产碱杆菌, 氰水解酶, 氰水合酶, 降解代谢途径, 产酶条件优化

Abstract:

A strain of Alcaligenes sp., DN25, isolated and preserved in the authors’ lab, shows a high cyanide-degrading activity and thus has a potential of industrial application.The cyanide-degrading routes were determined based on the analysis of degradation products and time curves.The experiments on optimization of culture conditions for the cyanide-degrading enzyme were carried out according to the characteristics of enzyme active site.The results demonstrated that the reaction was through a hydrolytic pathway co-functioned by a cyanide hydratase, an amidase and a cyanide hydrolase, among which cyanide hydrolase played a major role.Four sulfur-containing substances were added to the medium to test the effect on specific activity of culture.DL-cysteine was found to be beneficial for the enzyme production and DL-methionine for both enzyme production and cell yield, resulting in increased specific activity of culture of 1.26 U·ml^-1 and 2.10 U·ml^-1，when 0.8% DL-cysteine and DL-methionine was added into fermentation culture respectively.The experiments on optimization of liquid culture conditions in 1L shake flask gave the optimal conditions as 100ml of liquid loading, pH of 9.0, inoculation size of 8%, and 0.8% DL-methionine added into the medium.Under such conditions, the specific activity of culture could reach 2.18 U·ml^-1, increased by 3.46 fold compared to that before optimization.The cell growth curve had a similar trend to that of enzyme production and the stabilized phase could be reached within 40 h.The current work could lay the basis for further application and purification of the cyanide-degradating enzyme.

Key words: 产碱杆菌, 氰水解酶, 氰水合酶, 降解代谢途径, 产酶条件优化