5ZDN

The complex structure of FomD with CDP

Summary for 5ZDN

• Entry DOI: 10.2210/pdb5zdn/pdb
• Descriptor: FomD, CYTIDINE-5'-DIPHOSPHATE, MAGNESIUM ION, ... (5 entities in total)
• Functional Keywords: metalloprotein, hydrolase
• Biological source: Streptomyces fradiae (Streptomyces roseoflavus)
• Total number of polymer chains: 1
• Total formula weight: 24587.85

Authors

Sato, S.,Miyanaga, A.,Kudo, F.,Eguchi, T. (deposition date: 2018-02-23, release date: 2018-07-25, Last modification date: 2023-11-22)

Primary citation

Sato, S.,Miyanaga, A.,Kim, S.Y.,Kuzuyama, T.,Kudo, F.,Eguchi, T.
Biochemical and Structural Analysis of FomD That Catalyzes the Hydrolysis of Cytidylyl ( S)-2-Hydroxypropylphosphonate in Fosfomycin Biosynthesis.
Biochemistry, 57:4858-4866, 2018

PubMed Abstract: In fosfomycin biosynthesis, the hydrolysis of cytidylyl ( S)-2-hydroxypropylphosphonate [( S)-HPP-CMP] to afford ( S)-HPP is the only uncharacterized step. Because FomD is an uncharacterized protein with a DUF402 domain that is encoded in the fosfomycin biosynthetic gene cluster, FomD was hypothesized to be responsible for this reaction. In this study, FomD was found to hydrolyze ( S)-HPP-CMP to give ( S)-HPP and CMP efficiently in the presence of Mn or Co. FomD also hydrolyzed cytidylyl 2-hydroxyethylphosphonate (HEP-CMP), which is a biosynthetic intermediate before C-methylation. The k/ K value of FomD with ( S)-HPP-CMP was 10-fold greater than that with HEP-CMP, suggesting that FomD hydrolyzes ( S)-HPP-CMP rather than HEP-CMP in bacteria. The crystal structure of FomD showed that this protein adopts a barrel-like fold, which consists of a large twisted antiparallel β-sheet. This is a key structural feature of the DUF402 domain-containing proteins. Two metal cations are located between the FomD barrel and the two α-helices at the C-terminus and serve to presumably activate the phosphonate group of substrates for hydrolysis. Docking simulations with ( S)-HPP-CMP suggested that the methyl group at the C2 position of the HPP moiety is recognized by a hydrophobic interaction with Trp68. Further mutational analysis suggested that a conserved Tyr107 among the DUF402 domain family of proteins activates a water molecule to promote nucleophilic attack on the phosphorus atom of the phosphonate moiety. These findings provide mechanistic insights into the FomD reaction and lead to a complete understanding of the fosfomycin biosynthetic pathway in Streptomyces.

PubMed: 30010320
DOI: 10.1021/acs.biochem.8b00690

Experimental method

X-RAY DIFFRACTION (2.02 Å)