9O6A

CryoEM structure of EcKatG S-Trp105 at 2.22 Angstrom resolution revealing an asymmetric sulfur center in O=S-Trp

This is a non-PDB format compatible entry.

Summary for 9O6A

• Entry DOI: 10.2210/pdb9o6a/pdb
• EMDB information: 70168
• Descriptor: Catalase-peroxidase, PROTOPORPHYRIN IX CONTAINING FE (3 entities in total)
• Functional Keywords: met-tyr-trp cofactor, heme-dependent enzyme, non-canonical amino acid, oxidoreductase
• Biological source: Escherichia coli K-12
• Total number of polymer chains: 4
• Total formula weight: 326888.64

Authors

Duan, R.,Li, J.,Nathan, B.,Yang, X.,Liu, A. (deposition date: 2025-04-11, release date: 2026-04-22, Last modification date: 2026-08-19)

Primary citation

Duan, R.,Li, J.,Griffith, W.P.,Xu, Y.,Burrows, N.D.,Green, A.P.,Liu, A.
Single-atom substitution redirects KatG reactivity from cofactor biogenesis to stereoselective sulfoxidation.
Nat Commun, 17:-, 2026

PubMed Abstract: Protein-derived cofactors rely on precisely positioned heteroatoms to direct redox chemistry, yet isolating their individual contributions remains challenging. The indole N-H of tryptophan plays a central yet elusive role in biogenesis and function of the Met-Tyr-Trp (MYW) cofactor in catalase-peroxidase (KatG). Here, we use genetic code expansion to replace cofactor-forming Trp105 with thiotryptophan (S-Trp), enabling a single-heteroatom (N → S) substitution. Instead of forming the MYW crosslink, KatG bearing S-Trp105 undergoes site-specific monooxygenation to yield a chiral sulfoxide. HPLC-MS, circular dichroism, and FT-IR spectroscopy identify selective oxygen insertion at the sulfur, establishing enantioselective formation of an (S)-configured sulfoxide. A 2.22 Å cryo-EM structure visualizes the oxidized S-Trp105, revealing the S = O moiety orienting toward the iron and confirming the absence of crosslinking. The S-atom oxygenation is heme-dependent and proceeds via a two-electron oxygen-atom transfer, contrasting with the radical-mediated one-electron chemistry of native tryptophan. This redirection suppresses catalase activity by perturbing cofactor formation. These results show that a single-atom substitution reroutes the distal heme site from radical crosslinking to stereoselective sulfoxidation, uncovering a monooxygenase-like capability within KatG. This work highlights using noncanonical amino acids to achieve atomic-level control over reaction pathways and to interrogate cofactor biogenesis with unprecedented precision.

PubMed: 42288482
DOI: 10.1038/s41467-026-73579-y

Experimental method

ELECTRON MICROSCOPY (2.22 Å)