9DVH

A1 Tei + D-Hpg: Adenylation Domain 1 Core Construct from Teicoplanin Biosynthesis with D-4-Hydroxyphenylglycine

9DVH の概要

• エントリーDOI: 10.2210/pdb9dvh/pdb
• 分子名称: Non-ribosomal peptide synthetase, MbtH-like short polypeptide, (2R)-amino(4-hydroxyphenyl)ethanoic acid, ... (6 entities in total)
• 機能のキーワード: amp-binding enzyme, nrps, adenylation domain, ligase
• 由来する生物種: Actinoplanes teichomyceticus; 詳細
• タンパク質・核酸の鎖数: 4
• 化学式量合計: 100367.74

構造登録者

Hansen, M.H.,Cryle, M.J.,Ratnayake, M.S. (登録日: 2024-10-07, 公開日: 2025-08-13)

主引用文献

Ratnayake, M.S.,Jian, X.,Tailhades, J.,Challis, G.L.,Hansen, M.H.,Lewandowski, J.R.,Cryle, M.J.
Teicoplanin Nonribosomal Peptide Synthetase Is Unable to Incorporate Alpha-Ketoacid Building Blocks.
Biochemistry, 64:2039-2053, 2025

PubMed Abstract: Glycopeptide antibiotics (GPAs) are a vital class of nonribosomal peptides used as therapies of last resort to treat infections by multidrug-resistant bacteria. These peptide antibiotics are assembled by nonribosomal peptide synthetases (NRPSs), modular megasynthases central to the biosynthesis of a wide range of peptide natural products. The adenylation (A) domains of NRPSs are involved in the selection and activation of the amino acid building blocks forming these peptide natural products, with their subsequent loading onto a neighboring carrier protein for incorporation into the growing peptide chain. This makes A-domains the gatekeepers of specificity in nonribosomal peptide biosynthesis, with further studies needed to reveal how this specificity is enforced at all stages of catalysis. The first building block found in GPAs is diverse and can comprise an amino acid, a ketoacid, or mixtures of both, which suggests that the A-domains responsible for selecting these residues can also incorporate non-amino acid substrates. In this study, we explored the acceptance of such substrates by the initiation module of the teicoplanin NRPS. Our in vitro assays demonstrated that this A-domain possesses an unexpected preference for activating ketoacids over the native amino acid substrate l-Hpg. However, only (d/l)-Hpg and related amino acids were able to be loaded onto the neighboring carrier protein domain during the subsequent thioesterification step. We further characterized the structure of this A-domain from teicoplanin biosynthesis in complex with d-4-hydroxyphenylglycine (d-Hpg), which revealed alterations in the positioning of the substrate carboxylate that help explain the high levels of pyrophosphate release seen with this amino acid. In combination with extensive molecular dynamics simulations, these data suggest that ketoacid incorporation in GPA biosynthesis is likely performed after amino acid incorporation by the NRPS and highlight the importance of considering both activation and carrier protein loading reactions performed by an A-domain when investigating substrate selectivity in nonribosomal peptide biosynthesis.

PubMed: 40215197
DOI: 10.1021/acs.biochem.4c00770

実験手法

X-RAY DIFFRACTION (1.656 Å)