US EPA

7467666 

Journal Article 

Arginine deiminase uses an active-site cysteine in nucleophilic catalysis of L-arginine hydrolysis 

Lu, X; Galkin, A; Herzberg, O; Dunaway-Mariano, D 

2004 

Yes 

Journal of the American Chemical Society
ISSN: 0002-7863
EISSN: 1520-5126 

126 

17 

5374-5375 

English 

15113205 

10.1021/ja049543p 

Arginine deiminase (EC 3.5.3.6) catalyzes the hydrolysis of l-arginine to citrulline and ammonium ion, which is the first step of the microbial l-arginine degradation pathway. The deiminase conserves the active-site Cys-His-Asp motif found in several related enzymes that catalyze group-transfer reactions from the guanidinium center of arginine-containing substrates. For each of these enzymes, nucleophilic catalysis by the conserved Cys has been postulated but never tested. In this communication we report the results from rapid quench studies of single-turnover reactions carried out with recombinant Pseudomonas aeruginosa arginine deiminase and limiting [14C-1]l-arginine. The citrulline-formation and arginine-decay curves measured at 25 degrees C were fitted to yield apparent rate constants k = 3.6 +/- 0.1 s-1 and k = 4.2 +/- 0.1 s-1, respectively. The time course for the formation (k =13 s-1) and decay (k = 6.5 s-1) of 14C-labeled enzyme defined a kinetically competent intermediate. Under the same reaction conditions, the Cys406Ser mutant failed to form the 14C-labeled enzyme intermediate. These results, along with the recently reported enzyme X-ray structure (Galkin, A.; Kulakova, L.; Sarikaya, E.; Lim, K.; Howard, A.; Herzberg, O. J. Biol. Chem. 2004, 279, 14001-14008, evidence a reaction pathway in which l-arginine deimination proceeds via a covalent enzyme intermediate formed by ammonia displacement from the arginine guanidinum carbon by the active-site Cys406. 

arginine deiminase; cysteine; article; catalysis; energy yield; enzyme active site; enzyme structure; high performance liquid chromatography; hydrolysis; Arginine; Binding Sites; Catalysis; Chromatography, High Pressure Liquid; Cysteine; Hydrolases; Hydrolysis; Models, Molecular; Molecular Structure; Ornithine Carbamoyltransferase; Phosphotransferases (Carboxyl Group Acceptor); Protein Structure, Tertiary; Pseudomonas aeruginosa; Stereoisomerism