Jhonatan A. Hernandez-Valdes , Myrthe aan de Stegge , Jos Hermans , Johan Teunis , Rinke J. van Tatenhove-Pel , Bas Teusink , Herwig Bachmann , Oscar P. Kuipers
{"title":"利用微滴生物传感和选择系统增强乳酸乳球菌氨基酸的产生和分泌","authors":"Jhonatan A. Hernandez-Valdes , Myrthe aan de Stegge , Jos Hermans , Johan Teunis , Rinke J. van Tatenhove-Pel , Bas Teusink , Herwig Bachmann , Oscar P. Kuipers","doi":"10.1016/j.mec.2020.e00133","DOIUrl":null,"url":null,"abstract":"<div><p>Amino acids are attractive metabolites for the pharmaceutical and food industry field. On one hand, the construction of microbial cell factories for large-scale production aims to satisfy the demand for amino acids as bulk biochemical. On the other hand, amino acids enhance flavor formation in fermented foods. Concerning the latter, flavor formation in dairy products, such as cheese is associated with the presence of lactic acid bacteria (LAB). In particular, <em>Lactococcus lactis</em>, one of the most important LAB, is used as a starter culture in fermented foods. The proteolytic activity of some <em>L. lactis</em> strains results in peptides and amino acids, which are flavor compounds or flavor precursors. However, it is still a challenge to isolate bacterial cells with enhanced amino acid production and secretion activity. In this work, we developed a growth-based sensor strain to detect the essential amino acids isoleucine, leucine, valine, histidine and methionine. Amino acids are metabolites that can be secreted by some bacteria. Therefore, our biosensor allowed us to identify wild-type <em>L. lactis</em> strains that naturally secrete amino acids, by using co-cultures of the biosensor strain with potential amino acid producing strains. Subsequently, we used this biosensor in combination with a droplet-based screening approach, and isolated three mutated <em>L. lactis</em> IPLA838 strains with 5–10 fold increased amino acid-secretion compared to the wild type. Genome re-sequencing revealed mutations in genes encoding proteins that participate in peptide uptake and peptide degradation. We argue that an unbalance in the regulation of amino acid levels as a result of these gene mutations may drive the accumulation and secretion of these amino acids. This biosensing system tackles the problem of selection for overproduction of secreted molecules, which requires the coupling of the product to the producing cell in the droplets.</p></div>","PeriodicalId":18695,"journal":{"name":"Metabolic Engineering Communications","volume":null,"pages":null},"PeriodicalIF":3.7000,"publicationDate":"2020-12-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://sci-hub-pdf.com/10.1016/j.mec.2020.e00133","citationCount":"17","resultStr":"{\"title\":\"Enhancement of amino acid production and secretion by Lactococcus lactis using a droplet-based biosensing and selection system\",\"authors\":\"Jhonatan A. Hernandez-Valdes , Myrthe aan de Stegge , Jos Hermans , Johan Teunis , Rinke J. van Tatenhove-Pel , Bas Teusink , Herwig Bachmann , Oscar P. Kuipers\",\"doi\":\"10.1016/j.mec.2020.e00133\",\"DOIUrl\":null,\"url\":null,\"abstract\":\"<div><p>Amino acids are attractive metabolites for the pharmaceutical and food industry field. On one hand, the construction of microbial cell factories for large-scale production aims to satisfy the demand for amino acids as bulk biochemical. On the other hand, amino acids enhance flavor formation in fermented foods. Concerning the latter, flavor formation in dairy products, such as cheese is associated with the presence of lactic acid bacteria (LAB). In particular, <em>Lactococcus lactis</em>, one of the most important LAB, is used as a starter culture in fermented foods. The proteolytic activity of some <em>L. lactis</em> strains results in peptides and amino acids, which are flavor compounds or flavor precursors. However, it is still a challenge to isolate bacterial cells with enhanced amino acid production and secretion activity. In this work, we developed a growth-based sensor strain to detect the essential amino acids isoleucine, leucine, valine, histidine and methionine. Amino acids are metabolites that can be secreted by some bacteria. Therefore, our biosensor allowed us to identify wild-type <em>L. lactis</em> strains that naturally secrete amino acids, by using co-cultures of the biosensor strain with potential amino acid producing strains. Subsequently, we used this biosensor in combination with a droplet-based screening approach, and isolated three mutated <em>L. lactis</em> IPLA838 strains with 5–10 fold increased amino acid-secretion compared to the wild type. Genome re-sequencing revealed mutations in genes encoding proteins that participate in peptide uptake and peptide degradation. We argue that an unbalance in the regulation of amino acid levels as a result of these gene mutations may drive the accumulation and secretion of these amino acids. This biosensing system tackles the problem of selection for overproduction of secreted molecules, which requires the coupling of the product to the producing cell in the droplets.</p></div>\",\"PeriodicalId\":18695,\"journal\":{\"name\":\"Metabolic Engineering Communications\",\"volume\":null,\"pages\":null},\"PeriodicalIF\":3.7000,\"publicationDate\":\"2020-12-01\",\"publicationTypes\":\"Journal Article\",\"fieldsOfStudy\":null,\"isOpenAccess\":false,\"openAccessPdf\":\"https://sci-hub-pdf.com/10.1016/j.mec.2020.e00133\",\"citationCount\":\"17\",\"resultStr\":null,\"platform\":\"Semanticscholar\",\"paperid\":null,\"PeriodicalName\":\"Metabolic Engineering Communications\",\"FirstCategoryId\":\"1085\",\"ListUrlMain\":\"https://www.sciencedirect.com/science/article/pii/S2214030120300122\",\"RegionNum\":0,\"RegionCategory\":null,\"ArticlePicture\":[],\"TitleCN\":null,\"AbstractTextCN\":null,\"PMCID\":null,\"EPubDate\":\"\",\"PubModel\":\"\",\"JCR\":\"Q2\",\"JCRName\":\"BIOTECHNOLOGY & APPLIED MICROBIOLOGY\",\"Score\":null,\"Total\":0}","platform":"Semanticscholar","paperid":null,"PeriodicalName":"Metabolic Engineering Communications","FirstCategoryId":"1085","ListUrlMain":"https://www.sciencedirect.com/science/article/pii/S2214030120300122","RegionNum":0,"RegionCategory":null,"ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"Q2","JCRName":"BIOTECHNOLOGY & APPLIED MICROBIOLOGY","Score":null,"Total":0}
Enhancement of amino acid production and secretion by Lactococcus lactis using a droplet-based biosensing and selection system
Amino acids are attractive metabolites for the pharmaceutical and food industry field. On one hand, the construction of microbial cell factories for large-scale production aims to satisfy the demand for amino acids as bulk biochemical. On the other hand, amino acids enhance flavor formation in fermented foods. Concerning the latter, flavor formation in dairy products, such as cheese is associated with the presence of lactic acid bacteria (LAB). In particular, Lactococcus lactis, one of the most important LAB, is used as a starter culture in fermented foods. The proteolytic activity of some L. lactis strains results in peptides and amino acids, which are flavor compounds or flavor precursors. However, it is still a challenge to isolate bacterial cells with enhanced amino acid production and secretion activity. In this work, we developed a growth-based sensor strain to detect the essential amino acids isoleucine, leucine, valine, histidine and methionine. Amino acids are metabolites that can be secreted by some bacteria. Therefore, our biosensor allowed us to identify wild-type L. lactis strains that naturally secrete amino acids, by using co-cultures of the biosensor strain with potential amino acid producing strains. Subsequently, we used this biosensor in combination with a droplet-based screening approach, and isolated three mutated L. lactis IPLA838 strains with 5–10 fold increased amino acid-secretion compared to the wild type. Genome re-sequencing revealed mutations in genes encoding proteins that participate in peptide uptake and peptide degradation. We argue that an unbalance in the regulation of amino acid levels as a result of these gene mutations may drive the accumulation and secretion of these amino acids. This biosensing system tackles the problem of selection for overproduction of secreted molecules, which requires the coupling of the product to the producing cell in the droplets.
期刊介绍:
Metabolic Engineering Communications, a companion title to Metabolic Engineering (MBE), is devoted to publishing original research in the areas of metabolic engineering, synthetic biology, computational biology and systems biology for problems related to metabolism and the engineering of metabolism for the production of fuels, chemicals, and pharmaceuticals. The journal will carry articles on the design, construction, and analysis of biological systems ranging from pathway components to biological complexes and genomes (including genomic, analytical and bioinformatics methods) in suitable host cells to allow them to produce novel compounds of industrial and medical interest. Demonstrations of regulatory designs and synthetic circuits that alter the performance of biochemical pathways and cellular processes will also be presented. Metabolic Engineering Communications complements MBE by publishing articles that are either shorter than those published in the full journal, or which describe key elements of larger metabolic engineering efforts.