{"title":"Development and optimization of a modular two-fragment LacI switch for enhanced biosensor applications","authors":"","doi":"10.1007/s12257-024-00020-w","DOIUrl":null,"url":null,"abstract":"<h3>Abstract</h3> <p>Being able to perform modular design of artificial transcription factors is useful in bioengineering and synthetic biology, particularly in the development of biosensors and therapeutics. This study aimed to develop a two-fragment transcription factor system by splitting a lactose repressor (LacI). To fragment LacI, we screened potential split positions from transposon-based insertional libraries that we generated to identify those positions that did not disturb the intrinsic activity of LacI. The interaction of protein tags fused with fragments induces the reassembly of LacI and recovers the isopropyl-β-D-thiogalactoside-dependent regulatory function. The split LacI-based biosensor was implemented on an in vitro platform using a cell-free protein expression system to facilitate accurate analytical studies with high reproducibility. This versatile platform holds great potential to realize the rapid and simple detection of protein–protein interactions in cell-free systems; thus, it can be further extended to disease diagnosis, particularly at the point-of-care.</p>","PeriodicalId":8936,"journal":{"name":"Biotechnology and Bioprocess Engineering","volume":null,"pages":null},"PeriodicalIF":2.5000,"publicationDate":"2024-02-19","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":"0","resultStr":null,"platform":"Semanticscholar","paperid":null,"PeriodicalName":"Biotechnology and Bioprocess Engineering","FirstCategoryId":"5","ListUrlMain":"https://doi.org/10.1007/s12257-024-00020-w","RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"Q3","JCRName":"BIOTECHNOLOGY & APPLIED MICROBIOLOGY","Score":null,"Total":0}
引用次数: 0
Abstract
Being able to perform modular design of artificial transcription factors is useful in bioengineering and synthetic biology, particularly in the development of biosensors and therapeutics. This study aimed to develop a two-fragment transcription factor system by splitting a lactose repressor (LacI). To fragment LacI, we screened potential split positions from transposon-based insertional libraries that we generated to identify those positions that did not disturb the intrinsic activity of LacI. The interaction of protein tags fused with fragments induces the reassembly of LacI and recovers the isopropyl-β-D-thiogalactoside-dependent regulatory function. The split LacI-based biosensor was implemented on an in vitro platform using a cell-free protein expression system to facilitate accurate analytical studies with high reproducibility. This versatile platform holds great potential to realize the rapid and simple detection of protein–protein interactions in cell-free systems; thus, it can be further extended to disease diagnosis, particularly at the point-of-care.
期刊介绍：
Biotechnology and Bioprocess Engineering is an international bimonthly journal published by the Korean Society for Biotechnology and Bioengineering. BBE is devoted to the advancement in science and technology in the wide area of biotechnology, bioengineering, and (bio)medical engineering. This includes but is not limited to applied molecular and cell biology, engineered biocatalysis and biotransformation, metabolic engineering and systems biology, bioseparation and bioprocess engineering, cell culture technology, environmental and food biotechnology, pharmaceutics and biopharmaceutics, biomaterials engineering, nanobiotechnology, and biosensor and bioelectronics.
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