9JJB

Class 1 state of the GfsA KSQ-ancestralAT chimeric didomain in complex with the GfsA ACP domain

Summary for 9JJB

• Entry DOI: 10.2210/pdb9jjb/pdb
• EMDB information: 61522
• Descriptor: Polyketide synthase GfsA, Polyketide synthase, N-[2-(acetylamino)ethyl]-N~3~-[(2R)-2-hydroxy-3,3-dimethyl-4-(phosphonooxy)butanoyl]-beta-alaninamide (3 entities in total)
• Functional Keywords: decarboxylase, polyketide synthase, lyase
• Biological source: Streptomyces graminofaciens; More
• Total number of polymer chains: 4
• Total formula weight: 147622.29

Authors

Chisuga, T.,Liao, Z.,Adachi, N.,Kawasaki, M.,Moriya, T.,Senda, T.,Kudo, F.,Eguchi, T.,Miyanaga, A. (deposition date: 2024-09-13, release date: 2025-08-06, Last modification date: 2025-08-13)

Primary citation

Chisuga, T.,Takinami, S.,Liao, Z.,Karasawa, M.,Adachi, N.,Kawasaki, M.,Moriya, T.,Senda, T.,Terada, T.,Kudo, F.,Eguchi, T.,Nakano, S.,Ito, S.,Miyanaga, A.
Ancestral sequence reconstruction as a tool for structural analysis of modular polyketide synthases.
Nat Commun, 16:6847-6847, 2025

PubMed Abstract: Modular polyketide synthases (PKSs) are large multi-domain enzymes critical for the biosynthesis of polyketide antibiotics. However, challenges with structural analysis limits our mechanistic understanding of modular PKSs. In this report, we explore the potential of ancestral sequence reconstruction (ASR) for structure analysis of target proteins. As a model, we focus on the FD-891 PKS loading module composed of ketosynthase-like decarboxylase (KS), acyltransferase (AT) and acyl carrier protein (ACP) domains. We construct a KSAncAT chimeric didomain by replacing the native AT with an ancestral AT (AncAT) using ASR. After confirming that KSAncAT chimeric didomain retains similar enzymatic function to the native KSAT didomain, we successfully determine a high-resolution crystal structure of the KSAncAT chimeric didomain and cryo-EM structures of the KS-ACP complex. These cryo-EM structures, which could not be determined for the native protein, exemplify the utility of ASR to enable cryo-EM single-particle analysis. Our findings demonstrate that integrating ASR with structural analysis provides deeper mechanistic insight into modular PKSs. Furthermore, applying ASR to a partial region of the targeted multi-domain proteins could expand the potential of ASR and may serve as a valuable framework for investigating the structure and function of various multi-domain proteins.

PubMed: 40715098
DOI: 10.1038/s41467-025-62168-0

Experimental method

ELECTRON MICROSCOPY (2.68 Å)