title: Unusual hepatic mitochondrial arginase in an Indian air-breathing teleost, Heteropneustes fossilis: purification and characterization.
abstract: A functional urea cycle with both cytosolic (ARG I) and mitochondrial (ARG II) arginase activity is present in the liver of an ureogenic air-breathing teleost, Heteropneustes fossilis. Antibodies against mammalian ARG II showed no cross-reactivity with the H. fossilis ARG II. ARG II was purified to homogeneity from H. fossilis liver. Purified ARG II showed a native molecular mass of 96 kDa. SDS-PAGE showed a major band at 48 kDa. The native enzyme, therefore, appears to be a homodimer. The pI value of the enzyme was 7.5. The purified enzyme showed maximum activity at pH 10.5 and 55 °C. The K(m) of purified ARG II for l-arginine was 5.25±1.12 mM. L-Ornithine and N(ω)-hydroxy-L-arginine showed mixed inhibition with K(i) values 2.16±0.08 and 0.02±0.004 mM respectively. Mn(+2) and Co(+2) were effective activators of arginase activity. Antibody raised against purified H. fossilis ARG II did not cross-react with fish ARG I, and mammalian ARG I and ARG II. Western blot with the antibodies against purified H. fossilis hepatic ARG II showed cross reactivity with a 96 kDa band on native PAGE and a 48 kDa band on SDS-PAGE. The molecular, immunological and kinetic properties suggest uniqueness of the hepatic mitochondrial ARG II in H. fossilis.
authors.affiliation: Biochemical Adaptation Laboratory, Department of Zoology, Banaras Hindu University, Varanasi-221005, India. shilp.vastava@gmail.com
authors.forename: Shilpee
authors.initials: S
authors.lastname: Srivastava
authors.forename: B K
authors.initials: BK
authors.lastname: Ratha
mesh_terms: D000818:Animals; D001119:Arginase / Q000302:isolation & purification / Q000378:metabolism*; D001120:Arginine / Q000031:analogs & derivatives / Q000378:metabolism; D055162:Biocatalysis / Q000187:drug effects; D015153:Blotting, Western; D002397:Catfishes / Q000378:metabolism*; D003035:Cobalt / Q000494:pharmacology; D004591:Electrophoresis, Polyacrylamide Gel; D004789:Enzyme Activation / Q000187:drug effects; D029941:Fish Proteins / Q000302:isolation & purification / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D007194:India; D007525:Isoelectric Focusing; D007700:Kinetics; D008345:Manganese / Q000494:pharmacology; D008930:Mitochondria, Liver / Q000201:enzymology*; D008970:Molecular Weight; D009952:Ornithine / Q000378:metabolism; D013379:Substrate Specificity; D013696:Temperature
chemical_list: D029941:Fish Proteins; D003035:Cobalt; D008345:Manganese; C068309:N(omega)-hydroxyarginine; D001120:Arginine; D009952:Ornithine; D001119:Arginase