Purification and characterization of acidolysin, an acidic metalloprotease produced by Clostridium acetobutylicum ATCC 824.
Acidolysin an extracellular protease produced by Clostridium acetobutylicum ATCC 824 was purified to homogeneity by anion-exchange chromatography with a recovery of 91%. The enzyme was a monomeric protein with a molecular weight of 44,000 as estimated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and an acidic isoelectric point of 3.3. Acidolysin was very sensitive to metal-chelating agents and phosphoramidon and was unaffected by sulfhydryl reagents. It was shown to be a calcium- and zinc-containing protease. It exhibited optimal activity against Azocoll at pH 5 and 45 degrees C. It was stable at low pH and heat labile above 50 degrees C. It exhibited specificity toward peptide bonds formed by the amino group of hydrophobic amino acids (isoleucine, leucine, and phenylalanine) and its NH2-terminal amino acid sequence showed a high degree of similarity with that of Bacillus subtilis neutral metalloprotease A. Acidolysin is the first phosphoramidon-sensitive, acidic zinc metalloprotease reported.
Croux
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Département de Génie Biochimique et Alimentaire, UA-Centre National de la Recherche Scientifique no. 544, Institut National des Sciences Appliquées, Toulouse, France.
Paquet
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Goma
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Soucaille
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D000595:Amino Acid Sequence; D000596:Amino Acids / Q000032:analysis; D001391:Azo Compounds / Q000378:metabolism; D002118:Calcium / Q000032:analysis; D002852:Chromatography, Ion Exchange; D003013:Clostridium / Q000201:enzymology*; D003094:Collagen / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D004795:Enzyme Stability; D006863:Hydrogen-Ion Concentration; D006868:Hydrolysis; D007328:Insulin / Q000378:metabolism; D007526:Isoelectric Point; D008666:Metalloendopeptidases / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism; D008969:Molecular Sequence Data; D008970:Molecular Weight; D010455:Peptides / Q000378:metabolism; D012083:Renin / Q000378:metabolism; D013054:Spectrophotometry, Atomic; D013379:Substrate Specificity; D013696:Temperature; D015032:Zinc / Q000032:analysis
D000596:Amino Acids; D001391:Azo Compounds; D007328:Insulin; D010455:Peptides; C028470:Azocoll; D003094:Collagen; D012083:Renin; D008666:Metalloendopeptidases; C067991:acidolysin; D015032:Zinc; D002118:Calcium