Probing the role of the hyper-reactive histidine residue of arginase.
Rat liver arginase (arginase I) is potently inactivated by diethyl pyrocarbonate, with a second-order rate constant of 113M(-1)s(-1) for the inactivation process at pH 7.0, 25 degrees C. Partial protection from inactivation is provided by the product of the reaction, l-ornithine, while nearly complete protection is afforded by the inhibitor pair, l-ornithine and borate. The role of H141 has been probed by mutagenesis, chemical modulation, and X-ray diffraction. The hyper-reactivity of H141 towards diethyl pyrocarbonate can be explained by its proximity to E277. A proton shuttling role for H141 is supported by its conformational mobility observed among the known arginase structures. H141 is proposed to serve as an acid/base catalyst, deprotonating the metal-bridging water molecule to generate the metal-bridging hydroxide nucleophile, and by protonating the amino group of the product to facilitate its departure.
Colleluori
Diana M
DM
Department of Biochemistry, Temple University School of Medicine, Philadelphia, PA 19140, USA.
Reczkowski
Robert S
RS
Emig
Frances A
FA
Cama
Evis
E
Cox
J David
JD
Scolnick
Laura R
LR
Compher
Kevin
K
Jude
Kevin
K
Han
Shoufa
S
Viola
Ronald E
RE
Christianson
David W
DW
Ash
David E
DE
D000818:Animals; D001119:Arginase / Q000037:antagonists & inhibitors / Q000737:chemistry*; D001881:Borates / Q000737:chemistry; D018360:Crystallography, X-Ray; D004047:Diethyl Pyrocarbonate / Q000737:chemistry; D006639:Histidine / Q000737:chemistry*; D007700:Kinetics; D008958:Models, Molecular; D016297:Mutagenesis, Site-Directed; D009952:Ornithine / Q000737:chemistry; D011487:Protein Conformation; D051381:Rats
D001881:Borates; D006639:Histidine; D009952:Ornithine; D001119:Arginase; D004047:Diethyl Pyrocarbonate