Expression, purification, and characterization of human type II arginase.
Human type II arginase, which is extrahepatic and mitochondrial in location, catalyzes the hydrolysis of arginine to form ornithine and urea. While type I arginases function in the net production of urea for excretion of excess nitrogen, type II arginases are believed to function primarily in the net production of ornithine, a precursor of polyamines, glutamate, and proline. Type II arginases may also regulate nitric oxide biosynthesis by modulating arginine availability for nitric oxide synthase. Recombinant human type II arginase was expressed in Escherichia coli and purified to apparent homogeneity. The Km of arginine for type II arginase is approximately 4.8 mM at physiological pH. Type II arginase exists primarily as a trimer, although higher order oligomers were observed. Borate is a noncompetitive inhibitor of the enzyme, with a Kis of 0.32 mM and a Kii of 0.3 mM. Ornithine, a product of the reaction catalyzed by arginase and a potent inhibitor of type I arginase, is a poor inhibitor of the type II isozyme. The findings presented here indicate that isozyme-selectivity exists between type I and type II arginases for binding of substrate and products, as well as inhibitors. Therefore, inhibitors with greater isozyme-selectivity for type II arginase may be identified and utilized for the therapeutic treatment of smooth muscle disorders, such as erectile dysfunction.
Colleluori
D M
DM
Temple University School of Medicine, Department of Biochemistry, Philadelphia, Pennsylvania 19140, USA.
Morris
S M
SM
Ash
D E
DE
D001119:Arginase / Q000037:antagonists & inhibitors / Q000737:chemistry* / Q000235:genetics; D001120:Arginine / Q000378:metabolism / Q000494:pharmacology; D001667:Binding, Competitive / Q000187:drug effects; D002384:Catalysis / Q000187:drug effects; D002413:Cations, Divalent / Q000494:pharmacology; D002850:Chromatography, Gel; D002852:Chromatography, Ion Exchange; D018076:DNA, Complementary / Q000235:genetics; D004578:Electron Spin Resonance Spectroscopy; D004591:Electrophoresis, Polyacrylamide Gel; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004795:Enzyme Stability / Q000187:drug effects; D015870:Gene Expression; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007527:Isoenzymes / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000235:genetics; D008928:Mitochondria / Q000201:enzymology; D009952:Ornithine / Q000096:biosynthesis / Q000494:pharmacology; D016133:Polymerase Chain Reaction; D011994:Recombinant Proteins / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000235:genetics; D020539:Sequence Analysis, Protein; D014508:Urea / Q000378:metabolism
D002413:Cations, Divalent; D018076:DNA, Complementary; D004791:Enzyme Inhibitors; D007527:Isoenzymes; D011994:Recombinant Proteins; D014508:Urea; D001120:Arginine; D009952:Ornithine; D001119:Arginase