Yuanlin Liu , Yingying Zhu , Liping Qiao , Lumeng Yu , Qian Lin , Wei Xu , Wenli Zhang , Wanmeng Mu
{"title":"穆勒Simonsiella muelleri α2,6-唾液基转移酶能够在依赖质粒和不依赖质粒的工程大肠杆菌中高效地生物合成6 ' -唾液基乳糖","authors":"Yuanlin Liu , Yingying Zhu , Liping Qiao , Lumeng Yu , Qian Lin , Wei Xu , Wenli Zhang , Wanmeng Mu","doi":"10.1016/j.ijbiomac.2025.145718","DOIUrl":null,"url":null,"abstract":"<div><div>6′-Sialyllactose (6′-SL) is the most abundant sialylated human milk oligosaccharide in breast milk. Its health effects have been widely confirmed, and its biological production attracts increasing attention. The metabolic pathway of 6′-SL synthesis is clear, and the typical metabolic engineering strategies for its production improvement have been reported. However, there are not many <em>α</em>2,6-sialyltransferase (<em>α</em>2,6-SiaT) candidates reported. In this study, <em>α</em>2,6-SiaT from <em>Simonsiella muelleri</em> ATCC 29453 was reported to be effective in vivo 6′-SL productivity in engineered <em>Escherichia coli</em>. Based on this <em>α</em>2,6-SiaT, a two-plasmid <em>E. coli</em> BL21(DE3) was constructed to produce 6′-SL. After plasmid combination optimization and RBS engineering, which optimizes gene expression and enhances target product production by designing and modifying ribosome binding site (RBS) sequences, the titer was enhanced to 3.405 and 21.020 g/L in shake-flask and fed-batch cultivation, respectively. In addition, <em>E. coli</em> MG1655 was engineered to produce 6′-SL by genomic multiple copy integration of pathway genes, and the fed-batch cultivation generated 17.220 g/L of 6′-SL without the use of a plasmid. This study provided a new <em>α</em>2,6-SiaT candidate for microbial synthesis of 6′-SL.</div></div>","PeriodicalId":333,"journal":{"name":"International Journal of Biological Macromolecules","volume":"320 ","pages":"Article 145718"},"PeriodicalIF":8.5000,"publicationDate":"2025-07-08","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":"0","resultStr":"{\"title\":\"Simonsiella muelleri α2,6-sialyltransferase enables efficient biosynthesis of 6′-sialyllactose in both plasmid-dependent and plasmid-free engineered Escherichia coli\",\"authors\":\"Yuanlin Liu , Yingying Zhu , Liping Qiao , Lumeng Yu , Qian Lin , Wei Xu , Wenli Zhang , Wanmeng Mu\",\"doi\":\"10.1016/j.ijbiomac.2025.145718\",\"DOIUrl\":null,\"url\":null,\"abstract\":\"<div><div>6′-Sialyllactose (6′-SL) is the most abundant sialylated human milk oligosaccharide in breast milk. Its health effects have been widely confirmed, and its biological production attracts increasing attention. The metabolic pathway of 6′-SL synthesis is clear, and the typical metabolic engineering strategies for its production improvement have been reported. However, there are not many <em>α</em>2,6-sialyltransferase (<em>α</em>2,6-SiaT) candidates reported. In this study, <em>α</em>2,6-SiaT from <em>Simonsiella muelleri</em> ATCC 29453 was reported to be effective in vivo 6′-SL productivity in engineered <em>Escherichia coli</em>. Based on this <em>α</em>2,6-SiaT, a two-plasmid <em>E. coli</em> BL21(DE3) was constructed to produce 6′-SL. After plasmid combination optimization and RBS engineering, which optimizes gene expression and enhances target product production by designing and modifying ribosome binding site (RBS) sequences, the titer was enhanced to 3.405 and 21.020 g/L in shake-flask and fed-batch cultivation, respectively. In addition, <em>E. coli</em> MG1655 was engineered to produce 6′-SL by genomic multiple copy integration of pathway genes, and the fed-batch cultivation generated 17.220 g/L of 6′-SL without the use of a plasmid. This study provided a new <em>α</em>2,6-SiaT candidate for microbial synthesis of 6′-SL.</div></div>\",\"PeriodicalId\":333,\"journal\":{\"name\":\"International Journal of Biological Macromolecules\",\"volume\":\"320 \",\"pages\":\"Article 145718\"},\"PeriodicalIF\":8.5000,\"publicationDate\":\"2025-07-08\",\"publicationTypes\":\"Journal Article\",\"fieldsOfStudy\":null,\"isOpenAccess\":false,\"openAccessPdf\":\"\",\"citationCount\":\"0\",\"resultStr\":null,\"platform\":\"Semanticscholar\",\"paperid\":null,\"PeriodicalName\":\"International Journal of Biological Macromolecules\",\"FirstCategoryId\":\"92\",\"ListUrlMain\":\"https://www.sciencedirect.com/science/article/pii/S0141813025062737\",\"RegionNum\":1,\"RegionCategory\":\"化学\",\"ArticlePicture\":[],\"TitleCN\":null,\"AbstractTextCN\":null,\"PMCID\":null,\"EPubDate\":\"\",\"PubModel\":\"\",\"JCR\":\"Q1\",\"JCRName\":\"BIOCHEMISTRY & MOLECULAR BIOLOGY\",\"Score\":null,\"Total\":0}","platform":"Semanticscholar","paperid":null,"PeriodicalName":"International Journal of Biological Macromolecules","FirstCategoryId":"92","ListUrlMain":"https://www.sciencedirect.com/science/article/pii/S0141813025062737","RegionNum":1,"RegionCategory":"化学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"Q1","JCRName":"BIOCHEMISTRY & MOLECULAR BIOLOGY","Score":null,"Total":0}
Simonsiella muelleri α2,6-sialyltransferase enables efficient biosynthesis of 6′-sialyllactose in both plasmid-dependent and plasmid-free engineered Escherichia coli
6′-Sialyllactose (6′-SL) is the most abundant sialylated human milk oligosaccharide in breast milk. Its health effects have been widely confirmed, and its biological production attracts increasing attention. The metabolic pathway of 6′-SL synthesis is clear, and the typical metabolic engineering strategies for its production improvement have been reported. However, there are not many α2,6-sialyltransferase (α2,6-SiaT) candidates reported. In this study, α2,6-SiaT from Simonsiella muelleri ATCC 29453 was reported to be effective in vivo 6′-SL productivity in engineered Escherichia coli. Based on this α2,6-SiaT, a two-plasmid E. coli BL21(DE3) was constructed to produce 6′-SL. After plasmid combination optimization and RBS engineering, which optimizes gene expression and enhances target product production by designing and modifying ribosome binding site (RBS) sequences, the titer was enhanced to 3.405 and 21.020 g/L in shake-flask and fed-batch cultivation, respectively. In addition, E. coli MG1655 was engineered to produce 6′-SL by genomic multiple copy integration of pathway genes, and the fed-batch cultivation generated 17.220 g/L of 6′-SL without the use of a plasmid. This study provided a new α2,6-SiaT candidate for microbial synthesis of 6′-SL.
期刊介绍:
The International Journal of Biological Macromolecules is a well-established international journal dedicated to research on the chemical and biological aspects of natural macromolecules. Focusing on proteins, macromolecular carbohydrates, glycoproteins, proteoglycans, lignins, biological poly-acids, and nucleic acids, the journal presents the latest findings in molecular structure, properties, biological activities, interactions, modifications, and functional properties. Papers must offer new and novel insights, encompassing related model systems, structural conformational studies, theoretical developments, and analytical techniques. Each paper is required to primarily focus on at least one named biological macromolecule, reflected in the title, abstract, and text.