Preparation and properties of ornithine-oxo-acid aminotransferase of rat kidney comparison with the liver enzyme

Kalita, C. C.; Kerman, Jules D.; Strecker, Harold J.

doi:10.1016/0005-2744(76)90325-9

Cited by 25 publications

(10 citation statements)

References 32 publications

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“…It was hard to identify any molecular size diff e r e n c e with hepatic OAT (lane 3) on the electrophoregram. The molecular weight of the hepatic OAT was reported as 43-46 kDa by SDS-gel electrophoresis (Kalita et al, 1976;Lyons and Pitot, 1976;Sanada et al, 1976), and confirmed by the available primary structure in rat, to be 45,749 (Simmaco et al, 1986).…”

Section: Resultsmentioning

confidence: 83%

A variant of ornithine aminotransferase from mouse small intestine

Lim

Rho²,

Park³

et al. 1998

Exp Mol Med

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The ornithine aminotransferase (OAT) activity of mouse was found to be highest in the small intestine. The mitochondrial OAT from mouse small intestine was purified to homogeneity by the procedures including heat treatment, ammonium sulfate fractionation, octyl-S e p h a r o s e chromatography, and Sephadex G-150 gel filtration. Comparing to the amino acid sequence of mouse hepatic OAT, six N-terminal amino acid r e s i d u e s have been deleted in intestinal OAT. However, t h e subsequent sequence was identical with that of hepatic OAT. The molecular weights of both intestinal and hepatic OAT were estimated as 46 kDa by SDSgel electrophoresis and as 92 kDa by gel filtration, indicating that both native OATs are dimeric. Biochemical properties of intestinal OAT, such as molecular weight, pH optimum and K m values for Lornithine and -ketoglutarate, were similar to those of hepatic OAT. However, intestinal OAT was more labile than hepatic OAT to tryptic digestion. Keywords: ornithine aminotransferase; mouse; small intestine IntroductionOrnithine aminotransferase (OAT; L-ornithine:2-oxo-acid aminotransferase; EC 2.6.1.13) is a pyridoxal phosphaterequiring enzyme which catalyzes the reversible transamination of L-ornithine and a-ketoglutarate to glutamate and glutamic-γ-semialdehyde (its cyclized form being Δ 1 -pyrroline-5-carboxylate), and the latter product can be reversibly converted to no undeeline (Peraino and Pitot, 1963). OAT is expressed in nearly all mammalian tissues, including liver, kidney, brain, skeletal muscle, and eyes. OAT plays a role in arginine catabolism, proline biosynthesis, or de novo ornithine biosynthesis, depending on the tissue and the physiological circumstances (Mestichelli et al., 1979; Merril and Pitot,1983). Liver OAT is suggested to be involved in the ornithine synthesis for urea cycle while the kidney OAT participates in ornithine degradation (Herzfeld and Knox, 1968;Volpe et al. , 1969). Matsuzawa et al. (1994) suggested that the intestinal OAT may be involved in the ornithine supply to the liver, with the reversal of OAT reaction. In human, a genetic deficiency of OAT causes gyrate atrophy, an autosomal recessive degenerative disease of the choroid and retina of the eye that leads to blindness ( Valle et al., 1977;Kobayashi et al., 1995).OAT is known as a mitochondrial matrix enzyme and has been purified from various tissue sources, including rat liver, kidney, and brain (Matsuzawa et al., 1968;Sanada et al., 1970;Deshmukh, 1984) as well as human liver (Ohura et al., 1982). The mitochondrial OAT has been shown to be synthesized as a large precursor molecule with N-terminal leader peptide on cytoplasmic ribosome, which is then processed and becomes associated with the mitochondrion (Mueckler and Pitot, 1985). The amino acid sequence of OAT precursor protein was predicted from the nucleotide sequence of cDNA (Mueckler and Pitot, 1985;Inaga et al., 1986;Ramesh et al., 1986;Giometti et al., 1992). Recently it was reported that human kidney OAT had the same nucleotide sequenc...

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Section: Resultsmentioning

confidence: 83%

A variant of ornithine aminotransferase from mouse small intestine

Lim

Rho²,

Park³

et al. 1998

Exp Mol Med

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“…Although there is general consensus in the molecular mass of the polypeptides, which ranges over 43-45 kDa for OAT of different species and organs (Peraino et al, 1969;Kalita et al, 1976;Sanada et al, 1976), the holoenzyme is assumed to range over 90-280 kDa. Based on sedimentation equilibrium ultracentrifugation of rat liver OAT, MarkoviC-Housley et al (1987) reported a molecular mass of 256 kDa and suggested the enzyme to be composed of six subunits.…”

Section: Resultsmentioning

confidence: 99%

High‐resolution Electron Microscopic Studies on the Quaternary Structure of Ornithine Aminotransferase from Pig Kidney

Lünsdorf

Hecht

Tsai

1994

European Journal of Biochemistry

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The ornithine aminotransferase (OAT) from pig kidney has been studied on the basis of highresolution electron microscopy and the morphological appearance of the apoenzyme and holoenzyme have been examined. The quaternary structure of the OAT molecules in the presence of 5 mM pyridoxal 5'-phosphate could be established. The enzyme molecule appears to be built up of two morphological units, called M1. The native holoenzyme, termed morphological unit M2, measures . 10.9 nm in length and 5.8 nm in width and its molecular mass is approximately 168 kDa, based on electron microscopical calculations. Since the enzyme is composed of only one type of 45-kDa subunit, the holoenzyme is a homotetramer. Each M1 is composed of two subunits and, as seen in top-view projection, has an oval to triangular shape. Upon tilting to 40" the triangular shape changes into three distinct centers of mass. This morphological differentiation reflects the inner organization of M1, i.e. the shape of the individual subunit deviates from strictly globular proteins. This observation is compatible with the notion that the 45-kDa subunit consists of one large and one small domain. By tilting to 40", both large domains in M1 represent two of the three centers of mass, while the third center of mass is attributed to the superposition of both small domains. Thus, the four domains of both subunits in M1, in accordance with the triangular top-view projection, are quasi-tetrahedrally arranged. Since the change in shape of M1 upon tilting is only obvious in one of the two halves of the native OAT, it suggests that both morphological units of M2 are oriented asymmetrically relative to one another.

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“…1) (37). The nuclear encoded mitochondrial OAT has been isolated and purified from male rat kidney, characterized and visualized in male rat proximal tubules by immunohistochemistry (14,25). Finally, it is generally assumed that the genes encoding the enzymes that metabolize L-ornithine to L-citrulline and Lproline are expressed in the liver and the intestine but not in the rat kidney.…”

mentioning

confidence: 99%

“…In addition, to establish a metabolic link between AII and the enzymes that process in metabolizing L-arginine-derived ornithine, the distribution of OAT and AII was analyzed by Western blotting in whole kidneys, renal zones, and isolated mitochondria, whereas ODC mRNAs were localized by in situ hybridization. Finally, to prove that Lornithine could be metabolized to L-glutamate by OAT, Lornithine oxidation was analyzed by incubating dissected tubules with L- [1-14 C]ornithine or L-[U- 14 C]ornithine as sole substrates.…”

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confidence: 99%

Ornithine metabolism in male and female rat kidney: mitochondrial expression of ornithine aminotransferase and arginase II

Levillain

Hus-Citharel²,

Garvi³

et al. 2004

American Journal of Physiology-Renal Physiology

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.-In the kidney, L-ornithine is reabsorbed along the proximal convoluted tubule (PCT), transported by basolateral carriers, and produced by arginase II (AII). Here, the renal metabolic fate of L-ornithine was analyzed in male and female rats. Kidneys and renal zones were dissected and used for Western blot analysis, immunofluorescence, and electron microscopic studies. Ornithine aminotransferase (OAT) and AII were localized using specific antibodies. Ornithine oxidation was determined by incubating microdissected tubules with L-[1-14 C] or L-[U-14 C]ornithine in the presence or absence of energy-providing substrates. Ornithine decarboxylase (ODC) mRNAs were localized by in situ hybridization. The 48-kDa OAT protein was detected in male and female kidneys, but its level was fourfold higher in the latter. OAT relative distribution increased from the superficial cortex toward the outer medulla to reach its highest level. Almost all OAT protein was localized in cortical and medullary proximal straight tubules (CPST and OSPST, respectively). In proximal straight tubule (PST), AII protein distribution overlapped that of OAT. No gender difference in AII protein level was found. OAT and AII were colocalized within PST mitochondria. 14 C]ornithine demonstrated the complete oxidation of ornithine. In conclusion, the OAT gene was expressed more in female rat proximal tubules than in male. Because OAT and AII proteins overlapped in PST mitochondria, L-argininederived ornithine may be preferentially converted to L-glutamate, as proven by ornithine oxidation. However, the coexpression of ODC, glutamate decarboxylase, and glutamine synthetase in PST suggests that L-ornithine can also be metabolized to putrescine, GABA, and L-glutamine. The fate of L-ornithine may depend on the cellular context. L-ornithine; L-arginine; proximal tubules; isolated nephron segments; Western blot analysis; immunofluorescence; electron microscopy; mitochondria; ornithine decarboxylase THE AMINO ACID L-ORNITHINE is absent from food and is not a constituent of proteins; therefore, vertebrate organisms are dependent on its formation. In kidneys, four sources of Lornithine can be considered: 1) L-ornithine is present in the blood and then transported across the basolateral membranes of a variety of renal cells (36,40); 2) L-ornithine is filtered in glomeruli and reabsorbed by specific carriers located in the apical membrane of the proximal convoluted tubule (PCT) (43); 3) in PCT, the enzyme L-arginine:glycine amidinotransferase (GAT; EC 2.1.4.1) metabolizes L-arginine in the presence of L-glycine and produces guanidinoacetic acid and Lornithine (42); and 4) in the whole proximal straight tubule (PST) of several mammals including rats, the extrahepatic arginase II isoenzyme (AII; EC 3.5.3.1) hydrolyzes L-arginine into urea and L-ornithine (23).L-Ornithine is known to play a pivotal role in several metabolic pathways as precursor for L-proline, L-glutamate, L-glutamine, L-citrulline, and L-arginine biosynthesis. Interestingly, renal tubular cells contain...

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Preparation and properties of ornithine-oxo-acid aminotransferase of rat kidney comparison with the liver enzyme

Cited by 25 publications

References 32 publications

A variant of ornithine aminotransferase from mouse small intestine

A variant of ornithine aminotransferase from mouse small intestine

High‐resolution Electron Microscopic Studies on the Quaternary Structure of Ornithine Aminotransferase from Pig Kidney

Ornithine metabolism in male and female rat kidney: mitochondrial expression of ornithine aminotransferase and arginase II

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