Characterization of a Protease Apparently Involved in Processing of Pre-ornithine Transcarbamylase of Rat Liver

Overview

Journal Proc Natl Acad Sci U S A

Specialty Science

Date 1980 Dec 1

PMID 7012832

Citations 13

Authors

M Mori

S Miura

M TATIBANA

P P Cohen

Affiliations

Soon will be listed here.

Abstract

The precursor of rat liver ornithine transcarbamylase (ornithine carbamoyltransferase; carbamoylphosphate:L-ornithine carbamoyltransferase, EC 2.1.3.3) (pre-ornithine transcarbamylase), which was synthesized in a reticulocyte lysate cell-free system, was converted to an apparently mature form of the enzyme by isolated rat liver mitochondria. The proteolytic processing involved two steps: (i) conversion of pre-ornithine transcarbamylase (39,400 daltons) to a product of about 37,000 daltons and (ii) further conversion to the apparently mature form of the enzyme (36,00 daltons). When mitochondria were subfractionated by digitonin treatment followed by sonication of a mitoplast fraction, the proteolytic activity catalyzing the first step was recovered mainly in a matrix fraction. Some activity was found in an intermembrane space fraction. The enzyme activity in the matrix fraction has an optimal pH at about 7.5. The activity was inhibited almost completely by 2 mM leupeptin and partly by 2 mM antipain but not significantly by other microbial protease inhibitors or serine protease inhibitors. It was inhibited strongly by 2 mM EDTA, 2 mM ethylene glycol bis(beta-aminoethyl ether)-N,N,N',N'-tetraacetate, 2 mM p-chloromercuriphenylsulfonate, and 2 mM Hg(CH3COO)2 but not by N-ethylmaleimide or iodoacetamide. These results suggest that pre-ornithine transcarbamylase is first transported into the mitochondrial matrix and converted there to the mature form of the enzyme by a novel neutral protease(s).

Citing Articles

Revisiting trends on mitochondrial mega-channels for the import of proteins and nucleic acids.

Campo M, Peixoto P, Martinez-Caballero S J Bioenerg Biomembr. 2016; 49(1):75-99.

PMID: 27146409 DOI: 10.1007/s10863-016-9662-z.

The requirement of heat shock cognate 70 protein for mitochondrial import varies among precursor proteins and depends on precursor length.

Terada K, Ueda I, Ohtsuka K, Oda T, ICHIYAMA A, Mori M Mol Cell Biol. 1996; 16(11):6103-9.

PMID: 8887640 PMC: 231613. DOI: 10.1128/MCB.16.11.6103.

Liver mitochondria contain an ATP-dependent, vanadate-sensitive pathway for the degradation of proteins.

Desautels M, GOLDBERG A Proc Natl Acad Sci U S A. 1982; 79(6):1869-73.

PMID: 7043466 PMC: 346082. DOI: 10.1073/pnas.79.6.1869.

Synthesis, intracellular transport, and processing of the precursors for mitochondrial ornithine transcarbamylase and carbamoyl-phosphate synthetase I in isolated hepatocytes.

Mori M, Morita T, Ikeda F, Amaya Y, TATIBANA M, Cohen P Proc Natl Acad Sci U S A. 1981; 78(10):6056-60.

PMID: 6947214 PMC: 348976. DOI: 10.1073/pnas.78.10.6056.

Posttranslational uptake and processing of in vitro synthesized ornithine transcarbamoylase precursor by isolated rat liver mitochondria.

Conboy J, ROSENBERG L Proc Natl Acad Sci U S A. 1981; 78(5):3073-7.

PMID: 6942417 PMC: 319502. DOI: 10.1073/pnas.78.5.3073.

References

Sottocasa G, Kuylenstierna B, Ernster L, Bergstrand A . An electron-transport system associated with the outer membrane of liver mitochondria. A biochemical and morphological study. J Cell Biol. 1967; 32(2):415-38. PMC: 2107253. DOI: 10.1083/jcb.32.2.415. View

Schnaitman C, Greenawalt J . Enzymatic properties of the inner and outer membranes of rat liver mitochondria. J Cell Biol. 1968; 38(1):158-75. PMC: 2107463. DOI: 10.1083/jcb.38.1.158. View

Laemmli U . Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970; 227(5259):680-5. DOI: 10.1038/227680a0. View

Marshall M, Cohen P . Ornithine transcarbamylase from Streptococcus faecalis and bovine liver. I. Isolation and subunit structure. J Biol Chem. 1972; 247(6):1641-53. View

Gamble J, Lehninger A . Transport of ornithine and citrulline across the mitochondrial membrane. J Biol Chem. 1973; 248(2):610-8. View

Bonner W, Laskey R . A film detection method for tritium-labelled proteins and nucleic acids in polyacrylamide gels. Eur J Biochem. 1974; 46(1):83-8. DOI: 10.1111/j.1432-1033.1974.tb03599.x. View

Kessler S . Rapid isolation of antigens from cells with a staphylococcal protein A-antibody adsorbent: parameters of the interaction of antibody-antigen complexes with protein A. J Immunol. 1975; 115(6):1617-24. View

Clarke S . A major polypeptide component of rat liver mitochondria: carbamyl phosphate synthetase. J Biol Chem. 1976; 251(4):950-61. View

Pelham H, Jackson R . An efficient mRNA-dependent translation system from reticulocyte lysates. Eur J Biochem. 1976; 67(1):247-56. DOI: 10.1111/j.1432-1033.1976.tb10656.x. View

10.

Clarke S . The polypeptides of rat liver mitochondria: identification of a 36,000 dalton polypeptide as the subunit of ornithine transcarbamylase. Biochem Biophys Res Commun. 1976; 71(4):1118-24. DOI: 10.1016/0006-291x(76)90769-5. View

11.

Pierson D, Cox S, Gilbert B . Human ornithine transcarbamylase. Purification and characterization of the enzyme from normal liver and the liver of a Reye's syndrome patient. J Biol Chem. 1977; 252(18):6464-9. View

12.

Tsuji H, Hattori N, Yamamoto T, Kato K . The synthesis of rat liver lysosomes. I. Comparison of the microsomal, golgi, and lysosomal beta-glucuronidases. J Biochem. 1977; 82(3):619-36. DOI: 10.1093/oxfordjournals.jbchem.a131737. View

13.

Aoki Y . Crystallization and characterization of a new protease in mitochondria of bone marrow cells. J Biol Chem. 1978; 253(6):2026-32. View

14.

KALOUSEK F, Francois B, ROSENBERG L . Isolation and characterization of ornithine transcarbamylase from normal human liver. J Biol Chem. 1978; 253(11):3939-44. View

15.

Aoyagi T, Ishizuka M, Takeuchi T, Umezawa H . Enzyme inhibitors in relation to cancer therapy. Jpn J Antibiot. 1977; 30 Suppl:121-32. View

16.

Umezawa H . Recent advances in bioactive microbial secondary metabolites. Jpn J Antibiot. 1977; 30 Suppl:138-63. View

17.

Cote C, Solioz M, Schatz G . Biogenesis of the cytochrome bc1 complex of yeast mitochondria. A precursor form of the cytoplasmically made subunit V. J Biol Chem. 1979; 254(5):1437-9. View

18.

Maccecchini M, Rudin Y, Blobel G, Schatz G . Import of proteins into mitochondria: precursor forms of the extramitochondrially made F1-ATPase subunits in yeast. Proc Natl Acad Sci U S A. 1979; 76(1):343-7. PMC: 382935. DOI: 10.1073/pnas.76.1.343. View

19.

Michel R, Wachter E, Sebald W . Synthesis of a larger precursor for the proteolipid subunit of the mitochondrial ATPase complex of Neurospora crassa in a cell-free wheat germ system. FEBS Lett. 1979; 101(2):373-6. DOI: 10.1016/0014-5793(79)81047-9. View

20.

Chua N, Schmidt G . Transport of proteins into mitochondria and chloroplasts. J Cell Biol. 1979; 81(3):461-83. PMC: 2110399. DOI: 10.1083/jcb.81.3.461. View