Details
| Original language | English |
|---|---|
| Pages (from-to) | 5136-5144 |
| Number of pages | 9 |
| Journal | Journal of the American Chemical Society |
| Volume | 144 |
| Issue number | 11 |
| Early online date | 9 Mar 2022 |
| Publication status | Published - 23 Mar 2022 |
| Externally published | Yes |
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.
ASJC Scopus subject areas
- Chemical Engineering(all)
- Catalysis
- Chemistry(all)
- General Chemistry
- Biochemistry, Genetics and Molecular Biology(all)
- Biochemistry
- Chemical Engineering(all)
- Colloid and Surface Chemistry
Sustainable Development Goals
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In: Journal of the American Chemical Society, Vol. 144, No. 11, 23.03.2022, p. 5136-5144.
Research output: Contribution to journal › Article › Research › peer review
}
TY - JOUR
T1 - Total in Vitro Biosynthesis of the Thioamitide Thioholgamide and Investigation of the Pathway
AU - Sikandar, Asfandyar
AU - Lopatniuk, Maria
AU - Luzhetskyy, Andriy
AU - Müller, Rolf
AU - Koehnke, Jesko
N1 - Funding Information: J.K. thanks the BBSRC for support (BB/V016059/1). R.M. would like to acknowledge DFG (Leibniz Award: MU 1254/32-1).
PY - 2022/3/23
Y1 - 2022/3/23
N2 - 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.
AB - 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.
UR - http://www.scopus.com/inward/record.url?scp=85126571171&partnerID=8YFLogxK
U2 - 10.1021/jacs.2c00402
DO - 10.1021/jacs.2c00402
M3 - Article
C2 - 35263083
AN - SCOPUS:85126571171
VL - 144
SP - 5136
EP - 5144
JO - Journal of the American Chemical Society
JF - Journal of the American Chemical Society
SN - 0002-7863
IS - 11
ER -