{"title":"比较分子动力学方法指导糖基转移酶的定制,以满足合成应用的需要","authors":"","doi":"10.1039/d4gc01508h","DOIUrl":null,"url":null,"abstract":"<div><p>The properties of natural products can be significantly influenced by glycosylation, emphasizing the key role of glycosyltransferases (GTs) in this process. The pursuit of tailored GT catalysts to meet the demands of the glycosylation industry aligns with the principles of green chemistry. However, steering GT engineering towards the desired direction often requires substantial effort. Herein, we employ a comparative molecular dynamics approach to guide the engineering of GTs to alter their catalytic performances. Through comparing the structural flexibility and site-saturation mutagenesis of two GTs BarGT-1 with narrow substrate scope and BarGT-3 with wide substrate scope, the identified substitution K321P in C-loop 5 of BarGT-1 greatly expanded the substrate scope towards diverse pharmaceutically valuable substrates, thereby the catalytic efficiencies were remarkably improved (<em>e.g.</em>, 52- and 244-fold for 4′- and 6-hydroxyflavone, <em>k</em><sub>cat</sub>/<em>K</em><sub>M</sub>). Further, phylogenetic analysis demonstrated that lysine was a highly conserved residue in the GTs within BarGT-1 branch, and its key roles in regulating the substrate scope were validated through site-saturation mutagenesis in two novel GTs, BsyGT (K321F) and BgoGT (K322W). Moreover, the corresponding substitutions G325R/D in C-loop 5 of BarGT-3 also served as switches to regulate the substrate profile of BarGT-3. Finally, through the utilization of BarGT-1 and K321P, we successfully regulated the synthesis of valuable liquiritigenin glycosides with minor effort. The comparative molecular dynamics approach provides insights into the structural dynamics of GTs, optimizes enzymatic processes, and facilitates precise enzyme engineering, offering a directed synthetic paradigm to produce valuable glycosides.</p></div>","PeriodicalId":78,"journal":{"name":"Green Chemistry","volume":null,"pages":null},"PeriodicalIF":9.3000,"publicationDate":"2024-08-12","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://pubs.rsc.org/en/content/articlepdf/2024/gc/d4gc01508h?page=search","citationCount":"0","resultStr":"{\"title\":\"A comparative molecular dynamics approach guides the tailoring of glycosyltransferases to meet synthetic applications†\",\"authors\":\"\",\"doi\":\"10.1039/d4gc01508h\",\"DOIUrl\":null,\"url\":null,\"abstract\":\"<div><p>The properties of natural products can be significantly influenced by glycosylation, emphasizing the key role of glycosyltransferases (GTs) in this process. The pursuit of tailored GT catalysts to meet the demands of the glycosylation industry aligns with the principles of green chemistry. However, steering GT engineering towards the desired direction often requires substantial effort. Herein, we employ a comparative molecular dynamics approach to guide the engineering of GTs to alter their catalytic performances. Through comparing the structural flexibility and site-saturation mutagenesis of two GTs BarGT-1 with narrow substrate scope and BarGT-3 with wide substrate scope, the identified substitution K321P in C-loop 5 of BarGT-1 greatly expanded the substrate scope towards diverse pharmaceutically valuable substrates, thereby the catalytic efficiencies were remarkably improved (<em>e.g.</em>, 52- and 244-fold for 4′- and 6-hydroxyflavone, <em>k</em><sub>cat</sub>/<em>K</em><sub>M</sub>). Further, phylogenetic analysis demonstrated that lysine was a highly conserved residue in the GTs within BarGT-1 branch, and its key roles in regulating the substrate scope were validated through site-saturation mutagenesis in two novel GTs, BsyGT (K321F) and BgoGT (K322W). Moreover, the corresponding substitutions G325R/D in C-loop 5 of BarGT-3 also served as switches to regulate the substrate profile of BarGT-3. Finally, through the utilization of BarGT-1 and K321P, we successfully regulated the synthesis of valuable liquiritigenin glycosides with minor effort. The comparative molecular dynamics approach provides insights into the structural dynamics of GTs, optimizes enzymatic processes, and facilitates precise enzyme engineering, offering a directed synthetic paradigm to produce valuable glycosides.</p></div>\",\"PeriodicalId\":78,\"journal\":{\"name\":\"Green Chemistry\",\"volume\":null,\"pages\":null},\"PeriodicalIF\":9.3000,\"publicationDate\":\"2024-08-12\",\"publicationTypes\":\"Journal Article\",\"fieldsOfStudy\":null,\"isOpenAccess\":false,\"openAccessPdf\":\"https://pubs.rsc.org/en/content/articlepdf/2024/gc/d4gc01508h?page=search\",\"citationCount\":\"0\",\"resultStr\":null,\"platform\":\"Semanticscholar\",\"paperid\":null,\"PeriodicalName\":\"Green Chemistry\",\"FirstCategoryId\":\"92\",\"ListUrlMain\":\"https://www.sciencedirect.com/org/science/article/pii/S1463926224007003\",\"RegionNum\":1,\"RegionCategory\":\"化学\",\"ArticlePicture\":[],\"TitleCN\":null,\"AbstractTextCN\":null,\"PMCID\":null,\"EPubDate\":\"\",\"PubModel\":\"\",\"JCR\":\"Q1\",\"JCRName\":\"CHEMISTRY, MULTIDISCIPLINARY\",\"Score\":null,\"Total\":0}","platform":"Semanticscholar","paperid":null,"PeriodicalName":"Green Chemistry","FirstCategoryId":"92","ListUrlMain":"https://www.sciencedirect.com/org/science/article/pii/S1463926224007003","RegionNum":1,"RegionCategory":"化学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"Q1","JCRName":"CHEMISTRY, MULTIDISCIPLINARY","Score":null,"Total":0}
A comparative molecular dynamics approach guides the tailoring of glycosyltransferases to meet synthetic applications†
The properties of natural products can be significantly influenced by glycosylation, emphasizing the key role of glycosyltransferases (GTs) in this process. The pursuit of tailored GT catalysts to meet the demands of the glycosylation industry aligns with the principles of green chemistry. However, steering GT engineering towards the desired direction often requires substantial effort. Herein, we employ a comparative molecular dynamics approach to guide the engineering of GTs to alter their catalytic performances. Through comparing the structural flexibility and site-saturation mutagenesis of two GTs BarGT-1 with narrow substrate scope and BarGT-3 with wide substrate scope, the identified substitution K321P in C-loop 5 of BarGT-1 greatly expanded the substrate scope towards diverse pharmaceutically valuable substrates, thereby the catalytic efficiencies were remarkably improved (e.g., 52- and 244-fold for 4′- and 6-hydroxyflavone, kcat/KM). Further, phylogenetic analysis demonstrated that lysine was a highly conserved residue in the GTs within BarGT-1 branch, and its key roles in regulating the substrate scope were validated through site-saturation mutagenesis in two novel GTs, BsyGT (K321F) and BgoGT (K322W). Moreover, the corresponding substitutions G325R/D in C-loop 5 of BarGT-3 also served as switches to regulate the substrate profile of BarGT-3. Finally, through the utilization of BarGT-1 and K321P, we successfully regulated the synthesis of valuable liquiritigenin glycosides with minor effort. The comparative molecular dynamics approach provides insights into the structural dynamics of GTs, optimizes enzymatic processes, and facilitates precise enzyme engineering, offering a directed synthetic paradigm to produce valuable glycosides.
期刊介绍:
Green Chemistry is a journal that provides a unique forum for the publication of innovative research on the development of alternative green and sustainable technologies. The scope of Green Chemistry is based on the definition proposed by Anastas and Warner (Green Chemistry: Theory and Practice, P T Anastas and J C Warner, Oxford University Press, Oxford, 1998), which defines green chemistry as the utilisation of a set of principles that reduces or eliminates the use or generation of hazardous substances in the design, manufacture and application of chemical products. Green Chemistry aims to reduce the environmental impact of the chemical enterprise by developing a technology base that is inherently non-toxic to living things and the environment. The journal welcomes submissions on all aspects of research relating to this endeavor and publishes original and significant cutting-edge research that is likely to be of wide general appeal. For a work to be published, it must present a significant advance in green chemistry, including a comparison with existing methods and a demonstration of advantages over those methods.