不同来源S-腺苷甲硫氨酸合酶在大肠杆菌中的表达及催化应用  

Expression and catalytic application of S-adenosylmethionine synthase from different sources in Escherichia coli

作　　者：Hero Nmeri Godspower 乔郅钠 徐美娟[1] 饶志明[1] Hero Nmeri Godspower;QIAO Zhina;XU Meijuan;RAO Zhiming(School of Biotechnology,Jiangnan University,Wuxi 214122,China)

机构地区：[1]江南大学生物工程学院,江苏无锡214122

出　　处：《食品与发酵工业》2023年第2期26-33,共8页Food and Fermentation Industries

基　　金：国家自然科学基金(32071470;31770058)。

摘　　要：S-腺苷甲硫氨酸(S-adenosylmethionine,SAM)是一种重要的代谢中间体,对机体代谢活动的正常运行起重要作用,它是由底物L-甲硫氨酸和ATP在SAM合酶催化下生成的,目前主要应用于医药行业。该研究首先以蜡状芽孢杆菌Bacillus cereus和谷氨酸棒杆菌Corynebacterium glutamicum基因组为模板扩增出SAM合酶编码基因BcmetK和CgmetK,以pETDueT1质粒为载体,以大肠杆菌E.coli BL21(DE3)为底盘细胞,构建了SAM合成菌株E.coli BL21/pETDuet1-BcmetK和E.coli BL21/pETDuet1-CgmetK。其次,针对SAM合成菌株全细胞催化合成体系条件进行了优化,重组大肠杆菌E.coli BL21/pETDuet1-BcmetK在最优条件[L-甲硫氨酸浓度50 mmol/L、ATP浓度50 mmol/L、1.2%(体积分数)的OD_(600)值约9.0的细胞悬浮液、600 mmol/L pTSoNa、50 mmol/L MgSO_(4)、100 mmol/L K_(2)SO_(4)、45℃、pH 8.0]下,连续转化5 h,获得了464 mg/L的SAM。重组大肠杆菌E.coli BL21/pETDuet1-CgmetK在最优条件[L-甲硫氨酸浓度50 mmol/L、ATP浓度40 mmol/L、0.8%(体积分数)的OD_(600)值约9.0的细胞悬浮液、800 mmol/L pTSoNa、50 mmol/L MgSO 4、100 mmol/L K_(2)SO 4、45℃、pH 8.5]下,连续转化5 h,获得了528 mg/L的SAM,可见,谷氨酸棒杆菌来源的SAM合酶催化能力优于蜡状芽孢杆菌来源的,更有利于SAM的合成。该研究成功构建了SAM合成菌株,虽然产量水平不高,但为SAM的可持续生物合成提供了重要借鉴。Reactions that require the transfer of chemical functional groups are crucial to the survival and viability of living organisms.One such functional group is the methyl group.Methylation of genomic DNA is critical for the control of gene expression.The natural donor of the methyl group for such reaction is S-adenosylmethionine(SAM).Besides the methyl group,SAM is able to donate all groups surrounding its sulphur atom in metabolism.It is produced in almost all living organisms,except certain parasites,and can participate as intermediate in the synthesis of many other chemical compounds,including glutathione,spermine and spermidine,ethylene,N-acyl homoserine lactone,coenzyme Q10,creatine,melatonin,phosphatidylcholine,methylcobalamin,norepinephrine and carnitine.It has been used to treat inflammation,depression,liver disease,Alzheimer’s disease,liver disease,fibromyalgia,osteoarthritis,and colon cancer.Its many pharmacological applications have led to its continuously growing demand which means that without improved and sustainable production,the demand may not be satisfied.SAM is generated by the substrate L-methionine and ATP under the catalysis of SAM synthase.Heterologous gene expression has popularly been implemented in the improvement of microbial production of a variety of fine chemicals in industry.Whole-cell biotransformation is a promising alternative for the microbial production of important chemicals in contrast to classical fermentation.Whereas classical fermentation might be flawed with the demerit that while being hopeful of high productivity due to successful channelling of pathways and overexpression of key enzyme(s),often,the microbial chassis might convert some substrate into biomass if they are allowed to continue growing.This limitation could be overcome if cells are employed in the conversion of substrates to product(bioconversion)in their resting state.Bioconversion using whole-cells has attracted attention as an eco-friendly and sustainable method for the production of valuable compounds.Its

关 键 词：蜡状芽孢杆菌 谷氨酸棒杆菌 大肠杆菌 S-腺苷甲硫氨酸合酶 S-腺苷甲硫氨酸 全细胞催化 

分 类 号：Q78[生物学—分子生物学]