Total in Vitro Biosynthesis of the Thioamitide Thioholgamide and Investigation of the Pathway

verfasst von: Asfandyar Sikandar, Maria Lopatniuk, Andriy Luzhetskyy, Rolf Müller, Jesko Koehnke
Abstract: Thioholgamides are ribosomally synthesized and posttranslationally modified peptides (RiPPs), with potent activity against cancerous cell lines and an unprecedented structure. Despite being one of the most structurally and chemically complex RiPPs, very few biosynthetic steps have been elucidated. Here, we report the complete in vitro reconstitution of the biosynthetic pathway. We demonstrate that thioamidation is the first step and acts as a gatekeeper for downstream processing. Thr dehydration follows thioamidation, and our studies reveal that both these modifications require the formation of protein complexes─ThoH/I and ThoC/D. Harnessing the power of AlphaFold, we deduce that ThoD acts as a lyase and also proposes putative catalytic residues. ThoF catalyzes the oxidative decarboxylation of the terminal Cys, and the subsequent macrocyclization is facilitated by ThoE. This is followed by Ser dehydration, which is also carried out by ThoC/D. ThoG is responsible for histidine bis-N-methylation, which is a prerequisite for His β-hydroxylation─a modification carried out by ThoJ. The last step of the pathway is the removal of the leader peptide by ThoK to afford mature thioholgamide.
Externe Organisation(en): Helmholtz-Zentrum für Infektionsforschung GmbH (HZI)
Universität des Saarlandes
University of Glasgow
Typ: Artikel
Journal: Journal of the American Chemical Society
Band: 144
Seiten: 5136-5144
Anzahl der Seiten: 9
ISSN: 0002-7863
Publikationsdatum: 23.03.2022
Publikationsstatus: Veröffentlicht
Peer-reviewed: Ja
ASJC Scopus Sachgebiete: Katalyse, Allgemeine Chemie, Biochemie, Kolloid- und Oberflächenchemie
Ziele für nachhaltige Entwicklung: SDG 3 – Gute Gesundheit und Wohlergehen
Elektronische Version(en): http://hdl.handle.net/10033/623172 (Zugang: Offen)
https://doi.org/10.1021/jacs.2c00402 (Zugang: Geschlossen)