title: Purification and characterization of a novel zinc-proteinase from cultures of Aeromonas hydrophila.
abstract: While searching for an enzyme capable of breaking epsilon-(gamma-Glu)-Lys isopeptide bonds cross-linking protein chains, we purified a metallo-proteinase which mimics the action of an isopeptidase on the gamma-chain dimers of cross-linked fibrin. The enzyme is present in the growth medium of the bacterium Aeromonas hydrophila, isolated from the intestinal tract of the leech Hirudo medicinalis. It is a 19-kDa protein which specifically hydrolyzes the Gly-Ala peptide bond within the Gly-Gly-Ala sequence, located near the cross-link site in the gamma-chain dimer of fibrin. Substrate specificity studies with a number of synthetic peptides suggest that the enzyme prefers Gly-Gly or acetyl-Gly in the P2 and P1 positions, respectively (Schecter, I., and Berger, A. (1967) Biochem. Biophys. Res. Commun. 27, 157-162). Nonpolar amino acid residues seem to be favored in the P1' and P2' positions. The enzyme contains one atom of zinc and is inhibited by 1,10-phenanthroline, but not by EDTA. Iodoacetate, leupeptin, diisopropyl fluorophosphate, phenylmethylsulfonyl fluoride, pepstatin, and alpha 2-macroglobulin have no effect on enzyme activity. Disulfide reducing reagents, such as dithiothreitol or 2-mercaptoethanol, inactivate the enzyme completely. The partial amino-terminal sequence shows 46% identity with a zinc metallo-proteinase from a strain of Lysobacter enzymogenes and 69% identity with the LasA protein from Pseudomonas aeruginosa.
authors: Department of Biology, Haverford College, Pennsylvania 19041.
authors: A G
authors: AG
authors: Loewy
authors: U V
authors: UV
authors: Santer
authors: M
authors: M
authors: Wieczorek
authors: J K
authors: JK
authors: Blodgett
authors: S W
authors: SW
authors: Jones
authors: J C
authors: JC
authors: Cheronis
mesh_terms: D016980:Aeromonas hydrophila / Q000201:enzymology*; D000595:Amino Acid Sequence; D002851:Chromatography, High Pressure Liquid; D004591:Electrophoresis, Polyacrylamide Gel; D006801:Humans; D006863:Hydrogen-Ion Concentration; D008666:Metalloendopeptidases / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism; D008969:Molecular Sequence Data; D017386:Sequence Homology, Amino Acid; D013379:Substrate Specificity
chemical_list: D008666:Metalloendopeptidases; C044194:microbial metalloproteinases