Premium
Entrapment of Phosphoryl Intermediates by SAICAR Synthetase
Author(s) -
Ginder Nathaniel D,
Binkowski Daniel J.,
Chen Xiaoming,
Nix Jay C.,
Fromm Herbert J.,
Honzatko Richard B.
Publication year - 2008
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.22.2_supplement.233
Subject(s) - chemistry , nucleotide , stereochemistry , enzyme , biosynthesis , reaction intermediate , phosphorylation , crystallography , biochemistry , catalysis , gene
SAICAR synthetase catalyzes the eighth step in bacterial de novo purine nucleotide biosynthesis and is a target in the development of antimicrobial agents. The proposed enzyme‐mediated reaction proceeds by the transfer of the γ‐phosphoryl group of ATP to the carboxyl group of CAIR, followed by the attack of the α‐amino group of L‐aspartate on the carbonyl phosphate intermediate. Presented here are structures and positional isotope exchange (PIX) data that are consistent with a carbonyl phosphate of CAIR as a reaction intermediate. Enzyme from E. coli crystallized with IMP, ATP and Mg 2+ , has 1‐N‐phosphoryl IMP in its active site, as evidenced by X‐ray diffraction data to 1.5 Å resolution. Moreover under similar conditions, AICAR is N‐phosphorylated in crystal structures. In PIX kinetics, 18 O in the β,γ‐ bridging position of 18 O‐γ ATP moves to a terminal position of the β‐phosphoryl group only in the presence of enzyme and CAIR. PIX and structural data are consistent with the formation of a stable phosphoryl intermediate of CAIR.