Carrier Protein Interaction with Competing Adenylation and Epimerization Domains in a Nonribosomal Peptide Synthetase Analyzed by FRET.
Angew Chem Int Ed Engl
; 63(20): e202317753, 2024 05 13.
Article
em En
| MEDLINE
| ID: mdl-38488324
ABSTRACT
In multi-domain nonribosomal peptide synthetases (NRPSs) the order of domains and their catalytic specificities dictate the structure of the peptide product. Peptidyl-carrier proteins (PCPs) bind activated amino acids and channel elongating peptidyl intermediates along the protein template. To this end, fine-tuned interactions with the catalytic domains and large-scale PCP translocations are necessary. Despite crystal structure snapshots of several PCP-domain interactions, the conformational dynamics under catalytic conditions in solution remain poorly understood. We report a FRET reporter of gramicidin S synthetase 1 (GrsA; with A-PCP-E domains) to study for the first time the interaction between PCP and adenylation (A) domain in the presence of an epimerization (E) domain, a competing downstream partner for the PCP. Bulk FRET measurements showed that upon PCP aminoacylation a conformational shift towards PCP binding to the A domain occurs, indicating the E domain acts on its PCP substrate out of a disfavored conformational equilibrium. Furthermore, the A domain was found to preferably bind the D-Phe-S-Ppant-PCP stereoisomer, suggesting it helps in establishing the stereoisomeric mixture in favor of the D-aminoacyl moiety. These observations surprisingly show that the conformational logic can deviate from the order of domains and thus reveal new principles in the multi-domain interplay of NRPSs.
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Texto completo:
1
Coleções:
01-internacional
Base de dados:
MEDLINE
Assunto principal:
Peptídeo Sintases
/
Transferência Ressonante de Energia de Fluorescência
Idioma:
En
Revista:
Angew Chem Int Ed Engl
Ano de publicação:
2024
Tipo de documento:
Article
País de afiliação:
Alemanha