海藻酸钠/明胶协同固定化假单胞菌B-3酶法合成L-半胱氨酸  

Enzymatic Syntheses of L-cysteine by Sodium Alginate/Gelatin Co-immobilized Pseudomonas sp.B-3

作　　者：王普[1] 尹江峰[1] 何军邀[1] 梁法勇[1] 

机构地区：[1]浙江工业大学药学院,杭州310032

出　　处：《中国食品学报》2011年第2期34-41,共8页Journal of Chinese Institute Of Food Science and Technology

基　　金：浙江省重大科技攻关项目(No.2006C13001)

摘　　要：目的:研究海藻酸钠/明胶协同固定化假单胞菌B-3的制备条件,以及固定化细胞酶法转化DL-2-氨基-Δ2-噻唑啉-4-羧酸(DL-ATC)生物合成L-半胱氨酸的工艺条件。方法:比较8种不同的细胞固定化方法,优选海藻酸钠/明胶协同固定化为假单胞菌B-3最佳的固定化方法,并对凝胶组成,增殖活化时间和菌体包埋量等条件进行优化;通过向转化液中添加表面活性剂Tween-60,N-氨甲酰-L-半胱氨酸酰胺水解酶(L-NCC水解酶)激活剂Mn2+和L-半胱氨酸脱巯基酶抑制剂盐酸羟胺来提高L-半胱氨酸产量。结果:以增殖活化10h的固定化细胞为酶源,DL-ATC·3H2O质量浓度为20g/L,pH8.0,42℃酶促反应10h,产物L-半胱氨酸含量达9.18g/L,较游离细胞提高了29.0%,底物转化率达75.83%。固定化细胞重复使用4批次后相对转化率仍能维持在初始值的71.5%。结论:海藻酸钠/明胶协同包埋假单胞菌B-3细胞具有良好的生物转化活性;转化液中添加适量的Tween-60、Mn2+和盐酸羟胺能明显提高L-半胱氨酸产量。Objective： To investigate immobilization of Pseudomonas sp.B-3 by sodium alginate/gelatin mixed gel,and the biosynthesis of L-cysteine from DL-2-amino-△～2-thiazoline-4-carboxylic acid （DL-ATC） by immobilized cells.Methods： Suitable method for the immobilization of Pseudomonas sp.B-3 was selected by the comparison of eight immobilization methods.The influences of some key factors such as gel constitution,cells embedded and activation time on enzyme activity were optimized respectively.Tween-60,N-carbamyl-L-cysteine amidohydrolase （L-NCC hydrolase） activator of Mn2＋ and L-cysteine desulfhydrase inhibitor of hydroxylamine was added into reaction solution to improve L-cysteine productivity.Results： Sodium alginate/gelatin co-immobilization showed both the highest enzyme activity and best gel strength.After 10 h activation for immobilized cells,the bioconversion was conducted at pH 8.0 and 42 ℃ for 10 h,9.18 g·L-1 of L-cysteine was formed from 20 g·L-1 of DL-ATC·3H2O,with the molar conversion rate of 75.83%.An increase of 29.0% for L-cysteine production was obtained catalyzed by immobilized cells in comparison with resting cells.After reused for four times,the relative molar conversion rate of L-cysteine remained 71.5% of the initial value.Conclusion： Sodium alginate/gelatin embedding method is suitable for immobilization of Pseudomonas sp.B-3.L-cysteine production was enhanced by the addition of Tween-60,Mn2＋ and hydroxylamine hydrochloride in reaction solution.

关 键 词：DL-ATC L-半胱氨酸 假单胞菌B-3 细胞固定化 

分 类 号：TS202.3[轻工技术与工程—食品科学]