Exploring the Substrate Flexibility of GrsB Thioesterase Leads to the Structural Reassignment of a Gramicidin S Variant.

IF 2.6 4区 生物学 Q3 BIOCHEMISTRY & MOLECULAR BIOLOGY
ChemBioChem Pub Date : 2025-07-02 DOI:10.1002/cbic.202500412
Sho Konno, Tomoe Mizuguchi, Atsuko Suzuki, Miyu Tanaka, Fumihiro Ishikawa, Akihiro Taguchi, Atsuhiko Taniguchi, Genzoh Tanabe, Yoshio Hayashi
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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.

探索GrsB硫酯酶的底物灵活性导致Gramicidin S变体的结构重分配。
Gramicidin S (GS)是由两个五肽合成的环状十肽。gramicidin S合成酶B (GrsB)的c端硫酯酶(TE)结构域将前体五肽二聚并环化产生的线性十肽。最近,一种GS变体(GS- sa)通过前体定向生物合成被报道,其中一个单一的D-Phe被L-Ser(烯丙基)取代。为了了解GrsB-TE如何处理这些修饰的前体,我们使用合成的线性肽研究了其底物特异性。GrsB-TE环化的底物在6位含有l -丝氨酸(烯丙基),但在1位不含。然而,酶合成的GS- sa的HPLC保留时间与报道的GS变体不同。进一步的结构和功能分析,包括1H NMR、抗菌试验和圆二色光谱,表明所报道的GS-SA含有D-Ser(烯丙基)而不是L-Ser(烯丙基)。这些发现揭示了GrsB-TE中以前未被认识到的立体化学柔韧性,并支持了报道的GS变异的结构修正。
本文章由计算机程序翻译,如有差异,请以英文原文为准。
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来源期刊
ChemBioChem
ChemBioChem 生物-生化与分子生物学
CiteScore
6.10
自引率
3.10%
发文量
407
审稿时长
1 months
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).
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