Fang Yuan , Guoying Li , Zilong Li , Mingming Li , Xiaobo Liu , Haiquan Yang , Xiaobin Yu
{"title":"通过系统工程策略在莫巴拉链霉菌中高效合成转谷氨酰胺酶","authors":"Fang Yuan , Guoying Li , Zilong Li , Mingming Li , Xiaobo Liu , Haiquan Yang , Xiaobin Yu","doi":"10.1016/j.crfs.2024.100756","DOIUrl":null,"url":null,"abstract":"<div><p>Transglutaminases (TGases) have been widely used in food, pharmaceutical, biotechnology, and other industries because of their ability to catalyze deamidation, acyl transfer, and crosslinking reactions between Ƴ-carboxamide groups of peptides or protein-bound glutamine and the Ɛ-amino group of lysine. In this study, we demonstrated an efficient systematic engineering strategy to enhance the synthesis of TGase in a recombinant <em>Streptomyces mobaraensis</em> smL2020 strain in a 1000-L fermentor. Briefly, the enzymatic properties of the TGase TG<sub>L2020</sub> from <em>S</em>. <em>mobaraensis</em> smL2020 and TGase TG<sub>LD</sub> from <em>S</em>. <em>mobaraensis</em> smLD were compared to obtain the TGase TG<sub>LD</sub> with perfected characteristics for heterologous expression in a recombinant <em>S</em>. <em>mobaraensis</em> smL2020ΔTG without the gene <em>tg</em><sub><em>L 2020</em></sub>. Through multiple engineering strategies, including promoter engineering, optimizing the signal peptides and recombination sites, and increasing copies of the expression cassettes, the final TG<sub>LD</sub> activity in the recombinant <em>S. mobaraensis</em> smL2020ΔTG: (P<sub><em>L2020</em></sub>-<em>sp</em><sub><em>L2020</em></sub>-<em>protg</em><sub><em>LD</em></sub>-<em>tg</em><sub><em>LD</em></sub><em>)</em><sub><em>2</em></sub> (<em>tg</em><sub><em>L2020</em></sub> <em>and</em> BT1) reached 56.43 U/mL and 63.18 U/mL in shake flask and 1000-L fermentor, respectively, which was the highest reported to date. With the improvement of expression level, the application scope of TG<sub>LD</sub> in the food industry will continue to expand. Moreover, the genetic stability of the recombinant strain maintained at more than 20 generations. These findings proved the feasibility of multiple systematic engineering strategies in synthetic biology and provided an emerging solution to improve biosynthesis of industrial enzymes.</p></div>","PeriodicalId":10939,"journal":{"name":"Current Research in Food Science","volume":null,"pages":null},"PeriodicalIF":6.2000,"publicationDate":"2024-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.sciencedirect.com/science/article/pii/S2665927124000820/pdfft?md5=273219913f3f5eb02edd26be14542d45&pid=1-s2.0-S2665927124000820-main.pdf","citationCount":"0","resultStr":"{\"title\":\"Efficient biosynthesis of transglutaminase in Streptomyces mobaraensis via systematic engineering strategies\",\"authors\":\"Fang Yuan , Guoying Li , Zilong Li , Mingming Li , Xiaobo Liu , Haiquan Yang , Xiaobin Yu\",\"doi\":\"10.1016/j.crfs.2024.100756\",\"DOIUrl\":null,\"url\":null,\"abstract\":\"<div><p>Transglutaminases (TGases) have been widely used in food, pharmaceutical, biotechnology, and other industries because of their ability to catalyze deamidation, acyl transfer, and crosslinking reactions between Ƴ-carboxamide groups of peptides or protein-bound glutamine and the Ɛ-amino group of lysine. In this study, we demonstrated an efficient systematic engineering strategy to enhance the synthesis of TGase in a recombinant <em>Streptomyces mobaraensis</em> smL2020 strain in a 1000-L fermentor. Briefly, the enzymatic properties of the TGase TG<sub>L2020</sub> from <em>S</em>. <em>mobaraensis</em> smL2020 and TGase TG<sub>LD</sub> from <em>S</em>. <em>mobaraensis</em> smLD were compared to obtain the TGase TG<sub>LD</sub> with perfected characteristics for heterologous expression in a recombinant <em>S</em>. <em>mobaraensis</em> smL2020ΔTG without the gene <em>tg</em><sub><em>L 2020</em></sub>. Through multiple engineering strategies, including promoter engineering, optimizing the signal peptides and recombination sites, and increasing copies of the expression cassettes, the final TG<sub>LD</sub> activity in the recombinant <em>S. mobaraensis</em> smL2020ΔTG: (P<sub><em>L2020</em></sub>-<em>sp</em><sub><em>L2020</em></sub>-<em>protg</em><sub><em>LD</em></sub>-<em>tg</em><sub><em>LD</em></sub><em>)</em><sub><em>2</em></sub> (<em>tg</em><sub><em>L2020</em></sub> <em>and</em> BT1) reached 56.43 U/mL and 63.18 U/mL in shake flask and 1000-L fermentor, respectively, which was the highest reported to date. With the improvement of expression level, the application scope of TG<sub>LD</sub> in the food industry will continue to expand. Moreover, the genetic stability of the recombinant strain maintained at more than 20 generations. These findings proved the feasibility of multiple systematic engineering strategies in synthetic biology and provided an emerging solution to improve biosynthesis of industrial enzymes.</p></div>\",\"PeriodicalId\":10939,\"journal\":{\"name\":\"Current Research in Food Science\",\"volume\":null,\"pages\":null},\"PeriodicalIF\":6.2000,\"publicationDate\":\"2024-01-01\",\"publicationTypes\":\"Journal Article\",\"fieldsOfStudy\":null,\"isOpenAccess\":false,\"openAccessPdf\":\"https://www.sciencedirect.com/science/article/pii/S2665927124000820/pdfft?md5=273219913f3f5eb02edd26be14542d45&pid=1-s2.0-S2665927124000820-main.pdf\",\"citationCount\":\"0\",\"resultStr\":null,\"platform\":\"Semanticscholar\",\"paperid\":null,\"PeriodicalName\":\"Current Research in Food Science\",\"FirstCategoryId\":\"97\",\"ListUrlMain\":\"https://www.sciencedirect.com/science/article/pii/S2665927124000820\",\"RegionNum\":2,\"RegionCategory\":\"农林科学\",\"ArticlePicture\":[],\"TitleCN\":null,\"AbstractTextCN\":null,\"PMCID\":null,\"EPubDate\":\"\",\"PubModel\":\"\",\"JCR\":\"Q1\",\"JCRName\":\"FOOD SCIENCE & TECHNOLOGY\",\"Score\":null,\"Total\":0}","platform":"Semanticscholar","paperid":null,"PeriodicalName":"Current Research in Food Science","FirstCategoryId":"97","ListUrlMain":"https://www.sciencedirect.com/science/article/pii/S2665927124000820","RegionNum":2,"RegionCategory":"农林科学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"Q1","JCRName":"FOOD SCIENCE & TECHNOLOGY","Score":null,"Total":0}
Efficient biosynthesis of transglutaminase in Streptomyces mobaraensis via systematic engineering strategies
Transglutaminases (TGases) have been widely used in food, pharmaceutical, biotechnology, and other industries because of their ability to catalyze deamidation, acyl transfer, and crosslinking reactions between Ƴ-carboxamide groups of peptides or protein-bound glutamine and the Ɛ-amino group of lysine. In this study, we demonstrated an efficient systematic engineering strategy to enhance the synthesis of TGase in a recombinant Streptomyces mobaraensis smL2020 strain in a 1000-L fermentor. Briefly, the enzymatic properties of the TGase TGL2020 from S. mobaraensis smL2020 and TGase TGLD from S. mobaraensis smLD were compared to obtain the TGase TGLD with perfected characteristics for heterologous expression in a recombinant S. mobaraensis smL2020ΔTG without the gene tgL 2020. Through multiple engineering strategies, including promoter engineering, optimizing the signal peptides and recombination sites, and increasing copies of the expression cassettes, the final TGLD activity in the recombinant S. mobaraensis smL2020ΔTG: (PL2020-spL2020-protgLD-tgLD)2 (tgL2020and BT1) reached 56.43 U/mL and 63.18 U/mL in shake flask and 1000-L fermentor, respectively, which was the highest reported to date. With the improvement of expression level, the application scope of TGLD in the food industry will continue to expand. Moreover, the genetic stability of the recombinant strain maintained at more than 20 generations. These findings proved the feasibility of multiple systematic engineering strategies in synthetic biology and provided an emerging solution to improve biosynthesis of industrial enzymes.
期刊介绍:
Current Research in Food Science is an international peer-reviewed journal dedicated to advancing the breadth of knowledge in the field of food science. It serves as a platform for publishing original research articles and short communications that encompass a wide array of topics, including food chemistry, physics, microbiology, nutrition, nutraceuticals, process and package engineering, materials science, food sustainability, and food security. By covering these diverse areas, the journal aims to provide a comprehensive source of the latest scientific findings and technological advancements that are shaping the future of the food industry. The journal's scope is designed to address the multidisciplinary nature of food science, reflecting its commitment to promoting innovation and ensuring the safety and quality of the food supply.