Biomaterial Database

Purification of a liver alkaline protease which degrades oxidatively modified glutamine synthetase. Characterization as a high molecular weight cysteine proteinase. 1985

A J Rivett

A nonlysosomal alkaline protease which degrades the oxidatively modified form of Escherichia coli glutamine synthetase has been purified to apparent homogeneity from rat and mouse liver acetone powders. Its molecular weight was determined to be 300,000 by Sephacryl S-300 gel filtration but results of further studies using high pressure liquid chromatography gel filtration suggest a value of 650,000. Examination of the subunit structure by sodium dodecyl sulfate-polyacrylamide gel electrophoresis showed multiple bands of molecular weights between 22,000 and 34,000. The alkaline protease was inhibited by thiol reagents. Phenylmethylsulfonyl fluoride, aprotinin, leupeptin, antipain, and chymostatin partially inhibited the protease. The inhibition by phenylmethylsulfonyl fluoride was prevented by dithiothreitol, and alpha 1-antitrypsin and soybean trypsin inhibitor did not inhibit. No inhibition was observed with metalloprotease inhibitors. The alkaline protease is active over a broad range of pH with optimum activity for the degradation of oxidized glutamine synthetase around pH 9.0. Its activity is not stimulated by MgATP. A study of the products of insulin B chain degradation demonstrated major cleavage sites at Gln13-Ala14, Leu15-Tyr16, Cys(SO3H)19-Gly20, Gln4-His5, and Leu17-Val18. Based on its endopeptidase activity and its inhibitor specificity, the alkaline protease should be classified as a cysteine proteinase. It appears to be distinct from previously described proteinases and is likely involved in nonlysosomal mechanisms of intracellular protein turnover.

UI	MeSH Term	Description	Entries
D007328	Insulin	A 51-amino acid pancreatic hormone that plays a major role in the regulation of glucose metabolism, directly by suppressing endogenous glucose production (GLYCOGENOLYSIS; GLUCONEOGENESIS) and indirectly by suppressing GLUCAGON secretion and LIPOLYSIS. Native insulin is a globular protein comprised of a zinc-coordinated hexamer. Each insulin monomer containing two chains, A (21 residues) and B (30 residues), linked by two disulfide bonds. Insulin is used as a drug to control insulin-dependent diabetes mellitus (DIABETES MELLITUS, TYPE 1).	Iletin,Insulin A Chain,Insulin B Chain,Insulin, Regular,Novolin,Sodium Insulin,Soluble Insulin,Chain, Insulin B,Insulin, Sodium,Insulin, Soluble,Regular Insulin
D008099	Liver	A large lobed glandular organ in the abdomen of vertebrates that is responsible for detoxification, metabolism, synthesis and storage of various substances.	Livers
D008970	Molecular Weight	The sum of the weight of all the atoms in a molecule.	Molecular Weights,Weight, Molecular,Weights, Molecular
D010084	Oxidation-Reduction	A chemical reaction in which an electron is transferred from one molecule to another. The electron-donating molecule is the reducing agent or reductant; the electron-accepting molecule is the oxidizing agent or oxidant. Reducing and oxidizing agents function as conjugate reductant-oxidant pairs or redox pairs (Lehninger, Principles of Biochemistry, 1982, p471).	Redox,Oxidation Reduction
D010450	Endopeptidases	A subclass of PEPTIDE HYDROLASES that catalyze the internal cleavage of PEPTIDES or PROTEINS.	Endopeptidase,Peptide Peptidohydrolases
D011480	Protease Inhibitors	Compounds which inhibit or antagonize biosynthesis or actions of proteases (ENDOPEPTIDASES).	Antiprotease,Endopeptidase Inhibitor,Endopeptidase Inhibitors,Peptidase Inhibitor,Peptidase Inhibitors,Peptide Hydrolase Inhibitor,Peptide Hydrolase Inhibitors,Peptide Peptidohydrolase Inhibitor,Peptide Peptidohydrolase Inhibitors,Protease Antagonist,Protease Antagonists,Antiproteases,Protease Inhibitor,Antagonist, Protease,Antagonists, Protease,Hydrolase Inhibitor, Peptide,Hydrolase Inhibitors, Peptide,Inhibitor, Endopeptidase,Inhibitor, Peptidase,Inhibitor, Peptide Hydrolase,Inhibitor, Peptide Peptidohydrolase,Inhibitor, Protease,Inhibitors, Endopeptidase,Inhibitors, Peptidase,Inhibitors, Peptide Hydrolase,Inhibitors, Peptide Peptidohydrolase,Inhibitors, Protease,Peptidohydrolase Inhibitor, Peptide,Peptidohydrolase Inhibitors, Peptide
D002845	Chromatography	Techniques used to separate mixtures of substances based on differences in the relative affinities of the substances for mobile and stationary phases. A mobile phase (fluid or gas) passes through a column containing a stationary phase of porous solid or liquid coated on a solid support. Usage is both analytical for small amounts and preparative for bulk amounts.	Chromatographies
D003546	Cysteine Endopeptidases	ENDOPEPTIDASES which have a cysteine involved in the catalytic process. This group of enzymes is inactivated by CYSTEINE PROTEINASE INHIBITORS such as CYSTATINS and SULFHYDRYL REAGENTS.
D004591	Electrophoresis, Polyacrylamide Gel	Electrophoresis in which a polyacrylamide gel is used as the diffusion medium.	Polyacrylamide Gel Electrophoresis,SDS-PAGE,Sodium Dodecyl Sulfate-PAGE,Gel Electrophoresis, Polyacrylamide,SDS PAGE,Sodium Dodecyl Sulfate PAGE,Sodium Dodecyl Sulfate-PAGEs
D004926	Escherichia coli	A species of gram-negative, facultatively anaerobic, rod-shaped bacteria (GRAM-NEGATIVE FACULTATIVELY ANAEROBIC RODS) commonly found in the lower part of the intestine of warm-blooded animals. It is usually nonpathogenic, but some strains are known to produce DIARRHEA and pyogenic infections. Pathogenic strains (virotypes) are classified by their specific pathogenic mechanisms such as toxins (ENTEROTOXIGENIC ESCHERICHIA COLI), etc.	Alkalescens-Dispar Group,Bacillus coli,Bacterium coli,Bacterium coli commune,Diffusely Adherent Escherichia coli,E coli,EAggEC,Enteroaggregative Escherichia coli,Enterococcus coli,Diffusely Adherent E. coli,Enteroaggregative E. coli,Enteroinvasive E. coli,Enteroinvasive Escherichia coli