US EPA

8163654 

Journal Article 

Regiochemical and stereochemical course of the reaction catalyzed by the fosfomycin resistance protein, FosA 

Bernât, BA; Lauçhlin, LT; Armstrong, RN 

1998 

Yes 

FASEB Journal
ISSN: 0892-6638
EISSN: 1530-6860 

12 

8 

A1429-A1429 

English 

10.1021/jo980014b 

WOS:000083961500840 

The broad-spectrum antibiotic, (lR,2S)-epoxypropylphosphonic acid (subsequently called fosfomycin), blocks cell wall biosynthesis by irreversibly inhibiting uridine-5'-diphospho-/V-acetylD-glucosamine enolpyruvyl transferase. A plasmid from clinical isolates was shown to encode a 16 kDa polypeptide (FosA) that catalyzes the addition of glutathione (GSH) to the antibiotic, rendering it inactive. We recently established FosA to be a manganese metalloglutathione transferase that is related to other Fe!* and Mn2* métalloenzymes (Bernât, B. A., Laughlin, L. T. and Armstrong, R. N. (1997) Biochemistry 36, 3050-3055). The regiochemistry of the GSfosfomycin product, originally reported to be attack at the C-l of fosfomycin with 1-D NMR, has been confirmed by 2-D NMR. We have found cysleine can also be a substrate for the enzyme, although the kJK, value is -400-fold lower for cysteine than that for GSH. We have determined the stereochemistry of the thiolate addition by solving the crystal structure of the cysteine-fosfomycin adduct. Each asymmetric unit contains two cysteine-fosfomycin molecules, four ammonium ions and two waters. Using the a-carbon of the cysteine as a reference, the relative configuration of the molecule was determined. The reaction proceeds with inversion of configuration at the C-l position of fosfomycin. This result is consistent with a SN2 reaction mechanism. Supported by NIH gram AI 42756.