Adenosylmethionine (SAM) plays a vital role in the organisms’ metabolic activity as an important intermediate metabolite and has been widely used in pharmaceutical industry involving the therapies for liver, nervous systems, osteoarthritis and so on. Thus, production of SAM by enzyme transformation process exhibits potential industrial prospects. Codon optimization of the adenosylmethionine synthase gene sam2 derived from S. cerevisiae BY4741 was performed, and the gene was ligated to pET-28a(+) and pET-3b(+), resulting in recombinant plasmids pET-28a(+)-sam2 and pET-3b(+)-sam2, respectively, and was heterologous expressed in E. coli BL21(DE3) and E. coli Rosseta (DE3) respectively. Results showed that the recombinant strain E. coli BL21/ pET28a(+)-sam2 exhibited higher enzymatic activity up to 0.184 U/mL. Thus, the recombinant strain E. coli BL21/ pET-28a(+)-sam2 was chosen for the optimization of fermentation condition to further improve the enzymatic activity. Under optimal conditions, the enzyme activity of the recombinant adenosylmethionine synthase reached 0.245 U/mL, which was 34% higher than that before optimization.
MAO Zhiyi
,
TAN Xinyuan
,
CAO Rong
,
ZHANG Xiaojuan
,
FU Jing
,
XU Jianguo
,
XU Guoqiang
,
ZHANG Xiaomei
,
XU Zhenghong
. Heterologous expression of adenosylmethionine synthase in Escherichia coli and optimization of enzyme-producing conditions[J]. Food and Fermentation Industries, 2020
, 46(10)
: 8
-13
.
DOI: 10.13995/j.cnki.11-1802/ts.023219
[1] MATO J M,ALVAREZ L,ORTIZ P,et al.S-adenosylmethionine synthesis:Molecular mechanisms and clinical implications[J].Pharmacology Therapeutics,1997,73(3):265-280.
[2] ZHAO Weijun,SHI Feng,HANG Baojian,et al.The improvement of SAM accumulation by integrating the endogenous methionine adenosyltransferase gene SAM2 in genome of the industrial Saccharomyces cerevisiae strain[J].Applied Biochemistry and Biotechnology,2016,178(6):1 263-1 272.
[3] CARMEN GATCÍA-RUIZ,MORALES A,COLELL A,et al.Feeding S-adenosyl-L-methionine attenuates both ethanol-induced depletion of mitochondrial glutathione and mitochondrial dysfunction in periportal and perivenous rat hepatocytes[J].Hepatology,1995,21(1):207-214.
[4] CANTONI G L.The nature of the active methyl donor formed enzymatically from L-methionine and adenosinetriphosphate[J].Journal of the American Chemical Society,1952,74(11):2 942-2 943.
[5] 张巍巍,刘云顺,王颖,等.S-腺苷蛋氨酸在肝病患者中抗抑郁作用的研究进展[J].现代生物医学进展,2015,15(6):1 196-1 197.
[6] CHRISTOPH J,BERG T,CHENG J,et al.Intrahepatic cholestasis in common chronic liver diseases[J].European Journal of Clinical Investigation,2013,43(10):1 069-1 083.
[7] CARPENTER D J.St John's wort and S-adenosyl methionine as "natural" alternatives to conventional antidepressants in the era of the suicidality boxed warning:What is the evidence for clinically relevant benefit?[J].Alternative Medicine Review A Journal of Clinical Therapeutic,2011,16(1):17.
[8] MORELLI V,NAQUIN C,WEAVER V.Alternative therapies for traditional disease states:Osteoarthritis[J].American family physician,2003,67(2):339-344.
[9] 魏晓楠.S-腺苷甲硫氨酸(SAM)在大肠杆菌中的合成[D].福建:集美大学,2014.
[10] KAMARTHAPU V,RAGAMPETA S,RAO K V,et al.Engineered Pichia pastoris for enhanced production of S-adenosylmethionine[J].AMB Express,2013,3(1):40.
[11] RUAN Liying,LI Lu,ZOU Dian,et al.Metabolic engineering of Bacillus amyloliquefaciens for enhanced production of S-adenosylmethionine by coupling of an engineered S-adenosylmethionine pathway and the tricarboxylic acid cycle[J].Biotechnology for Biofuels, 2019,12(1):211
[12] 王杰鹏.发酵法生产S-腺苷蛋氨酸前体蛋氨酸补加策略[J].生物工程学报,2008,24(10):1 824-1 827.
[13] MARKHAM G D,DEPARASIS J,GATMAITAN J.The sequence of metK,the structural gene for S-adenosylmethionine synthetase in Escherichia coli[J].Journal of Biological Chemistry,1985,259(23):14 505-14 507.
[14] 刘珞.高产S-腺苷蛋氨酸合成酶基因工程菌的构建[J].北京化工大学学报(自然科学版),2015,40(2):70-73.
[15] HU Xiaoqing,CHU Ju,ZHANG Siliang,et al.A novel feeding strategy during the production phase for enhancing the enzymatic synthesis of S-adenosyl-L-methionine by methylotrophic Pichia pastoris[J].Enzyme and Microbial Technology,2007,40(4):669-674.
[16] DIPPE M,BRANDT W,ROST H,et al.Rationally engineered variants of S-adenosylmethionine (SAM) synthase:Reduced product inhibition and synthesis of artificial cofactor homologues[J].Chem Commun,2015,51(17):3 637-3 640.
[17] KANAI M,MIZUNUMA M,FUJII T,et al.A genetic method to enhance the accumulation of S-adenosylmethionine in yeast[J].Applied Microbiology and Biotechnology,2017,101(4):1 351-1 357.
[18] DIPPE M, BRRANDT W,ROST H,et. Rationally engineered variants of S-adenosylmethionine (SAM) synthase: Reduced product inhibition and synthesis of artificial cofactor homologues[J].Chemical Communications,2019,51(17):3 637-3 640.
[19] 安胜欣.酿酒酵母S-腺苷甲硫氨酸合成酶基因的克隆及表达研究[D].北京:北京化工大学,2006.
[20] RONG Jingjing,DIAO Zhenyu,ZHOU Guohua.Research progress on E.coli expression system[J].Pharmaceutical Biotechnology,2005.
[21] 吴梧桐.生物制药工艺学(第二版)[M].北京:中国医药科技出版社,2006.
[22] DUAN Xuguo,CHEN Jian,WU Jing.Optimization of pullulanase production in Escherichia coli by regulation of process conditions and supplement with natural osmolytes[J].Bioresour Technol,2013,46(10):79-85.
