A high molecular weight metalloendoprotease from the cytosol of mammalian cells.
In red cell lysates, three soluble proteases hydrolyze insulin at pH 8.5. One of these enzymes was purified to homogeneity by conventional chromatographic techniques. It appears to be a metalloprotease since it is inhibited by EDTA, o-phenanthroline, and 8-hydroxyquinoline, the metal-depleted enzyme can be reactivated by micromolar levels of Zn2+, Co2+, or Mn2+, and it is not inhibited by reagents specific for carboxyl, serine or thiol proteases. This enzyme has an apparent molecular weight of 300,000 +/- 25,000, and electrophoresis in sodium dodecyl sulfate indicates a single band with an Mr = 115,000 +/- 10,000. End group analysis and automated Edman degradation of the products of proteolysis showed that it is an endoprotease which cleaves on the NH2-terminal side of large hydrophobic amino acids. Although various small polypeptides with Mr = 2300-3500 are hydrolyzed (e.g. insulin chains, glucagon, and calcitonin), a variety of larger proteins are not degraded (e.g. casein and globin). The latter proteins, however, are converted to substrates for the metalloprotease by digestion with the ATP-stimulated endoprotease from erythrocytes. Thus, the metalloprotease may play a role in the ATP-dependent pathway for degrading proteins with abnormal structures and could account in part for the o-phenanthroline sensitivity of this process. A similar enzyme is found in humans, rabbits, and rats and is cytosolic in all tissues which have been examined including erythrocytes, reticulocytes, liver, kidney, brain, and skeletal muscle.
D002850:Chromatography, Gel; D003600:Cytosol / Q000201:enzymology; D004492:Edetic Acid / Q000494:pharmacology; D010450:Endopeptidases / Q000302:isolation & purification*; D004912:Erythrocytes / Q000201:enzymology*; D005934:Glucagon / Q000378:metabolism; D006801:Humans; D006868:Hydrolysis; D007328:Insulin / Q000097:blood; D046911:Macromolecular Substances; D008666:Metalloendopeptidases; D008970:Molecular Weight
D007328:Insulin; D046911:Macromolecular Substances; D005934:Glucagon; D004492:Edetic Acid; D010450:Endopeptidases; D008666:Metalloendopeptidases