{"title":"Exploring the Substrate Flexibility of GrsB Thioesterase Leads to the Structural Reassignment of a Gramicidin S Variant.","authors":"Sho Konno, Tomoe Mizuguchi, Atsuko Suzuki, Miyu Tanaka, Fumihiro Ishikawa, Akihiro Taguchi, Atsuhiko Taniguchi, Genzoh Tanabe, Yoshio Hayashi","doi":"10.1002/cbic.202500412","DOIUrl":null,"url":null,"abstract":"<p><p>Gramicidin S (GS) is a cyclic decapeptide derived from two pentapeptides. The C-terminal thioesterase (TE) domain of gramicidin S synthetase B (GrsB) dimerizes precursor pentapeptides and cyclizes the resulting linear decapeptide. Recently, a GS variant (GS-SA), in which a single D-Phe was replaced by L-Ser(Allyl), was reported via precursor-directed biosynthesis in a native GS producer. To understand how GrsB-TE processes such modified precursors, we investigated its substrate specificity using synthetic linear peptides. GrsB-TE cyclized a substrate containing L-Ser(Allyl) at position 6 but not at position 1. However, the enzymatically synthesized GS-SA showed a different HPLC retention time than that of the reported GS variant. Further structural and functional analyses, including 1H NMR, antimicrobial assays, and circular dichroism spectroscopy, revealed that the reported GS-SA contained D-Ser(Allyl) rather than L-Ser(Allyl). These findings reveal a previously unrecognized stereochemical flexibility in GrsB-TE and support the structural revision of the reported GS variant.</p>","PeriodicalId":140,"journal":{"name":"ChemBioChem","volume":" ","pages":"e202500412"},"PeriodicalIF":2.6000,"publicationDate":"2025-07-02","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":"0","resultStr":null,"platform":"Semanticscholar","paperid":null,"PeriodicalName":"ChemBioChem","FirstCategoryId":"99","ListUrlMain":"https://doi.org/10.1002/cbic.202500412","RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"Q3","JCRName":"BIOCHEMISTRY & MOLECULAR BIOLOGY","Score":null,"Total":0}
引用次数: 0
Abstract
Gramicidin S (GS) is a cyclic decapeptide derived from two pentapeptides. The C-terminal thioesterase (TE) domain of gramicidin S synthetase B (GrsB) dimerizes precursor pentapeptides and cyclizes the resulting linear decapeptide. Recently, a GS variant (GS-SA), in which a single D-Phe was replaced by L-Ser(Allyl), was reported via precursor-directed biosynthesis in a native GS producer. To understand how GrsB-TE processes such modified precursors, we investigated its substrate specificity using synthetic linear peptides. GrsB-TE cyclized a substrate containing L-Ser(Allyl) at position 6 but not at position 1. However, the enzymatically synthesized GS-SA showed a different HPLC retention time than that of the reported GS variant. Further structural and functional analyses, including 1H NMR, antimicrobial assays, and circular dichroism spectroscopy, revealed that the reported GS-SA contained D-Ser(Allyl) rather than L-Ser(Allyl). These findings reveal a previously unrecognized stereochemical flexibility in GrsB-TE and support the structural revision of the reported GS variant.
期刊介绍:
ChemBioChem (Impact Factor 2018: 2.641) publishes important breakthroughs across all areas at the interface of chemistry and biology, including the fields of chemical biology, bioorganic chemistry, bioinorganic chemistry, synthetic biology, biocatalysis, bionanotechnology, and biomaterials. It is published on behalf of Chemistry Europe, an association of 16 European chemical societies, and supported by the Asian Chemical Editorial Society (ACES).