title: [Isolation and characteristics of Bacillus megaterium metalloproteinase].
abstract: Stepwise application of affinity chromatography on bacitracin-silochrome, gel filtration on Acrylex P-10, rechromatography on bacitracin-Sepharose 4B and gel filtration on Sephadex G-15, a homogeneous metalloproteinase (M(r) = 35,000 Da) has been isolated from the cultural filtrate of B. megaterium strain 599. The amino acid composition and N-terminal sequence (20 amino acids) of the enzyme have been determined. The proteinase is not inhibited by diisopropyl-fluorophosphate, is inhibited by o-phenanthroline, EDTA, and Zn2+, and is activated by Co2+. The enzyme has a peak activity at 60-65 degrees C. The maximum of the enzymatic activity after hydrolysis of synthetic substrates is at pH 6.5-7.0. The enzyme is stable at pH 7.0-9.0 and retains its stability at 45-60 C for several hours. In acid media the enzyme undergoes irreversible inactivation. The dependence of kcat/Km on pH points to the involvement of an ionogenic group with pKa 7.5 in the catalytic act, most probably of the imidazole group of histidine. The metalloproteinase hydrolyzes synthetic peptide substrates at the bonds formed by the amino groups of hydrophobic amino acids-Phe, Leu, Ile and Val.
authors.forename: I P
authors.initials: IP
authors.lastname: Morozova
authors.forename: G G
authors.initials: GG
authors.lastname: Chestukhina
authors.forename: M E
authors.initials: ME
authors.lastname: Bormatova
authors.forename: M Iu
authors.initials: MIu
authors.lastname: Gololobov
authors.forename: N M
authors.initials: NM
authors.lastname: Ivanova
authors.forename: E N
authors.initials: EN
authors.lastname: Lysogorskaia
authors.forename: I Iu
authors.initials: IIu
authors.lastname: Filippova
authors.forename: O M
authors.initials: OM
authors.lastname: Khodova
authors.forename: E A
authors.initials: EA
authors.lastname: Timokhina
authors.forename: V M
authors.initials: VM
authors.lastname: Stepanov
mesh_terms: D000595:Amino Acid Sequence; D001410:Bacillus megaterium / Q000201:enzymology*; D002846:Chromatography, Affinity; D002850:Chromatography, Gel; D004795:Enzyme Stability; D006863:Hydrogen-Ion Concentration; D006868:Hydrolysis; D008666:Metalloendopeptidases / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism; D008969:Molecular Sequence Data; D017386:Sequence Homology, Amino Acid; D013379:Substrate Specificity
chemical_list: D008666:Metalloendopeptidases
vernacular_title: Vydelenie i kharakteristika metalloproteinazy Bacillus megaterium.