Fosfomycin Biosynthesis via Transient Cytidylylation of 2‑Hydroxyethylphosphonate by the Bifunctional Fom1 Enzyme

• Published in: ACS chemical biology Vol. 12; no. 8; pp. 2209 - 2215
• Main Authors: Cho, Su-Hee, Kim, Seung-Young, Tomita, Takeo, Shiraishi, Taro, Park, Jin-Soo, Sato, Shusuke, Kudo, Fumitaka, Eguchi, Tadashi, Funa, Nobutaka, Nishiyama, Makoto, Kuzuyama, Tomohisa
• Format: Journal Article
• Language: English
• Published: United States American Chemical Society 18-08-2017
• Subjects: Catalytic Domain; Crystallization; Cytidine Monophosphate - chemistry; Cytidine Monophosphate - metabolism; Fosfomycin - biosynthesis; Fosfomycin - chemistry; Models, Biological; Models, Molecular; Organophosphonates - chemistry; Organophosphonates - metabolism; Protein Domains; Substrate Specificity
• ISSN: 1554-8929; 1554-8937
• DOI: 10.1021/acschembio.7b00419

Fosfomycin is a wide-spectrum phosphonate antibiotic that is used clinically to treat cystitis, tympanitis, etc. Its biosynthesis starts with the formation of a carbon–phosphorus bond catalyzed by the phosphoenolpyruvate phosphomutase Fom1. We identified an additional cytidylyltransferase (CyTase) domain at the Fom1 N-terminus in addition to the phosphoenolpyruvate phosphomutase domain at the Fom1 C-terminus. Here, we demonstrate that Fom1 is bifunctional and that the Fom1 CyTase domain catalyzes the cytidylylation of the 2-hydroxyethylphosphonate (HEP) intermediate to produce cytidylyl-HEP. On the basis of this new function of Fom1, we propose a revised fosfomycin biosynthetic pathway that involves the transient CMP-conjugated intermediate. The identification of a biosynthetic mechanism via such transient cytidylylation of a biosynthetic intermediate fundamentally advances the understanding of phosphonate biosynthesis in nature. The crystal structure of the cytidylyl-HEP-bound CyTase domain provides a basis for the substrate specificity and reveals unique catalytic elements not found in other members of the CyTase family.