Organic compounds -- part of the class 532-570 series – Organic compounds – Carbohydrates or derivatives
Patent
1993-09-10
1996-06-04
Wax, Robert A.
Organic compounds -- part of the class 532-570 series
Organic compounds
Carbohydrates or derivatives
435195, 435212, 435213, 435220, 435 691, 4353201, 536 221, 536 231, 536 237, C12N 914, C12N 948, C12N 952, C07H 1900
Patent
active
055233902
ABSTRACT:
Provide herein is a substantially pure gingipain-1 preparation, gingipain-1 being characterized as having an apparent molecular mass of 50 kDa as estimated by sodium dodecyl sulfate polyacrylamide gel electrophoresis and an apparent molecular mass of 44 kDa as estimated by gel filtration chromatography, said gingipain-1 having amidolytic and proteolytic activity for cleavage after arginine residues and having no amidolytic and/or proteolytic activity for cleavage after lysine residues, wherein the amidolytic and/or proteolytic activity is inhibited by cysteine protease group-specific inhibitors including iodoacetamide, iodoacetic acid, N-ethylmaleimide, leupeptin, antipain, trans-epoxysuccinyl-L-leucylamido-(4-guanidino)butane, TLCK, TPCK, p-aminobenzamidine, N-chlorosuccinamide, and chelating agents including EDTA and EGTA, wherein the amidolytic and/or proteolytic activity of said gingipain-1 is not sensitive to inhibition by human cystatin C, .alpha.2-macroglobulin, .alpha.1-proteinase inhibitor, antithrombin III, .alpha.2-antiplasmin, serine protease group-specific inhibitors including diisopropylfluorophosphate, phenylmethyl sulfonylfluoride and 3,4-diisochlorocoumarin, and wherein the amidolytic and/or proteolytic activities of gingipain-1 are stabilized by Ca.sup.2+ and wherein the amidolytic and/or proteolytic activities of said gingipain-1 are stimulated by glycine-containing peptides and glycine analogues, and methods for preparation. As specifically exemplified, Arg-gingipain-1 and Arg-gingipain-2 are purified from Porphyromonas gingivalis, and the mature Arg-gingipain-2 has an amino acid sequence as given in SEQ ID NO:5 from amino acid 1 through amino acid 510. Also provided are nucleic acid sequences encoding Arg-gingipain-2. The nucleotide coding sequence of the mature Arg-gingipain-2 is given in SEQ ID NO:4, from nucleotide 1630 through nucleotide 3105.
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Barr Philip J.
Pavloff Nadine
Potempa Jan S.
Travis James
Kim Hyosuk
University of Georgia Research Foundation Inc.
Wax Robert A.
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