Probing the role of the hyper-reactive histidine residue of arginase |
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Authors: | Colleluori Diana M Reczkowski Robert S Emig Frances A Cama Evis Cox J David Scolnick Laura R Compher Kevin Jude Kevin Han Shoufa Viola Ronald E Christianson David W Ash David E |
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Institution: | Department of Biochemistry, Temple University School of Medicine, Philadelphia, PA 19140, USA. |
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Abstract: | 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. |
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Keywords: | Arginase Chemical modification Chemical modulation Mutagenesis Diethyl pyrocarbonate Bimetallic hydrolases |
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