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Protein Engineering of Pasteurella multocida alpha 2,3-Sialyltransferase with Reduced alpha 2,3-Sialidase Activity and Application in Synthesis of 3 '-Sialyllactose

文献类型：期刊论文

作者Yang, Rui1,2; Gong, Mengge2,3; Jiao, Siming2; Han, Juntian2; Feng, Cui2; Pei, Meishan3; Zhou, Zhongkai1; Du, Yuguang2; Li, Jianjun2
刊名CATALYSTS
出版日期2022-06-01
卷号12期号:6页码:11
关键词sialyltransferases Pasteurella multocida alpha 2,3-sialyltransferase alpha 2,3-sialidase kinetic characterization 3 '-sialyllactose
DOI10.3390/catal12060579
英文摘要Sialyltransferases are key enzymes for the production of sialosides. The versatility of Pasteurella multocida alpha 2,3-sialyltransferase 1 (PmST1) causes difficulties in the efficient synthesis of alpha 2,3-linked sialylatetd compounds, especial its alpha 2,3-sialidase activity. In the current study, the alpha 2,3-sialidase activity of PmST1 was further reduced by rational design-based protein engineering. Three double mutants PMG1 (M144D/R313Y), PMG2 (M144D/R313H) and PMG3 (M144D/R313N) were designed and constructed using M144D as the template and kinetically investigated. In comparison with M144D, the alpha 2,3-sialyltransferase activity of PMG2 was enhanced by 1.4-fold, while its alpha 2,3-sialidase activity was reduced by 4-fold. Two PMG2-based triple mutants PMG2-1 (M144D/R313H/T265S) and PMG2-2 (M144D/R313H/E271F) were then designed, generated and characterized. Compared with PMG2, triple mutants showed slightly improved alpha 2,3-sialyltransferase activity, but their alpha 2,3-sialidase activities were increased by 2.1-2.9 fold. In summary, PMG2 was used for preparative-scale production of 3 '-SL (3 '-sialyllactose) with a yield of >95%. These new PmST1 mutants could be potentially utilized for efficient synthesis of alpha 2,3-linked sialosides. This work provides a guide to designing and constructing efficient sialyltransferases.
WOS关键词ALPHA/BETA-GALACTOSIDE ALPHA-2,3-SIALYLTRANSFERASE ; SIALIC-ACID ; CHEMOENZYMATIC SYNTHESIS ; NEISSERIA-MENINGITIDIS ; ESCHERICHIA-COLI ; SIALYLTRANSFERASE ; EXPRESSION ; CLONING ; IDENTIFICATION
资助项目National Natural Science Foundation of China[21877114]
WOS研究方向Chemistry
语种英语
出版者MDPI
WOS记录号WOS:000816401300001
资助机构National Natural Science Foundation of China
源URL[http://ir.ipe.ac.cn/handle/122111/54027]  
专题中国科学院过程工程研究所
通讯作者Zhou, Zhongkai; Du, Yuguang; Li, Jianjun
作者单位1.Tianjin Univ Sci & Technol, Key Lab Food Nutr & Safety, Minist Educ, Tianjin 300457, Peoples R China
2.Chinese Acad Sci, Inst Proc Engn, Natl Key Lab Biochem Engn, Natl Engn Res Ctr Biotechnol Beijing, Beijing 100190, Peoples R China
3.Univ Jinan, Sch Chem & Chem Engn, Jinan 250022, Peoples R China
推荐引用方式
GB/T 7714		 Yang, Rui,Gong, Mengge,Jiao, Siming,et al. Protein Engineering of Pasteurella multocida alpha 2,3-Sialyltransferase with Reduced alpha 2,3-Sialidase Activity and Application in Synthesis of 3 '-Sialyllactose[J]. CATALYSTS,2022,12(6):11.
APA Yang, Rui.,Gong, Mengge.,Jiao, Siming.,Han, Juntian.,Feng, Cui.,...&Li, Jianjun.(2022).Protein Engineering of Pasteurella multocida alpha 2,3-Sialyltransferase with Reduced alpha 2,3-Sialidase Activity and Application in Synthesis of 3 '-Sialyllactose.CATALYSTS,12(6),11.
MLA Yang, Rui,et al."Protein Engineering of Pasteurella multocida alpha 2,3-Sialyltransferase with Reduced alpha 2,3-Sialidase Activity and Application in Synthesis of 3 '-Sialyllactose".CATALYSTS 12.6(2022):11.

入库方式： OAI收割

来源：过程工程研究所

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