固定化重组细胞催化非天然二肽合成

Synthesis of Non-Natural Dipeptides Catalyzed by Immobilized Recombinant Cells

采用溴化十六烷基三甲基氯化铵(CTAB)处理增加细胞通透性;处理后细胞以聚乙烯醇为载体包埋,然后用2%(体积分数)戊二醛交联制得固定化重组细胞。选取有代表性的6种游离L-氨基酸为酰基受体,以D-苯甘氨酰胺为酰基供体、固定化A.faecalis PGA重组细胞为催化剂,进行了非天然二肽合成研究。结果显示,A.faecalis PGA催化反应速率较快,除D-Phenylglyl-L-glycine二肽开始时反应速率稍慢(液相得率20%)外,其余5种二肽的合成在反应15 min时液相得率均超过50%,应用潜力较好。批反应结果表明,固定化酶活相对较稳定,可以重复使用,且底物稳定性优于E.coli PGA。

Recombinant cells were permeabilized with CTAB to increase the enzyme activity.Treated cells were entrapped by polyvinyl alcohol and then crosslinked with 2%（volume fraction） glutaraldehyde to increase the stability.With the immobilized recombinant A.faecalis PGA cells as catalyst,D-phenylglycine amine as the acyl donor,and six kinds of typical L-amino acids as the acyl receptor,six non-natural dipeptides were synthesized.The results indicated that the reaction catalyzed by A.faecalis PGA was double-quick,and the HPLC yield of the synthetic dipeptides exceeded 50% in 15 min except D-phenylglyl-L-glycine（only 20%）.In the batch reaction,the enzyme activity of the immobilized cells was relatively steady and they could be reused successively.Furthermore,it was found that A.faecalis PGA had better stability than E.coli PGA in higher substrate concentration.Based on these observations,A.faecalis PGA has the better potential to be used in synthesis of dipeptides.