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Preliminary data on purified kallikrein from rat kidney
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Conference Abstracts
Vol . 16, No . 5
Of the four rat urinary kallikrein fractions noted by Nustad and Pierce (1), the moat abundant was isolated using Whatman DE-32 cellulose and affinity chromatography . The latter involved the use of Aprotinin (TrasylolR) bound by the amide method to Bio-Gel P-200 . Using SDS-polyacrylamide gel electrophoresis, molecular weights of 37,000, 31,000 and 32,000 were estimated respectively for human, rabbit and rat urinary kallikreina . The respective esterase activities were 30, 67, and 1030 esterase unite/mg (apectrophotometric assay using benzoyl-L-arginine ethyl ester) . Conspicuous differences in the amino acid composition of the kallikrein were found. 1.
R. NUSTAD and J .V . PIERCE, Biochemistry Z3 2312-2319 (1974) . PRELIMINARY DATA ON PURIFIED RALLIKREIN FROM RAT KIDNEY G. Porcelli, R.H . Croxatto, G.B . Marini-Hettalo and M . Di Iorio
Instituto di Chimica delle Facoltà di Medicine dell'Univeraità Cattolica, Rome Centro dells Chimica dei Recettori del C .N .R ., Rome Laboratorio de Fiaiologa della Univereidad Catolica, Santiago, Chile A highly purified kallikrein from rat kidney was obtained by a multi-step purification procedure including gel filtration on Sephadex 6-100, chromatography on Whatman CM-32 and DE 32 celluloses and Sephadex G-100 superfine. Molecular weight of 40,000 was estimated for the purified enzyme by sodium dodecyl sulfate polyacrylamide disc gel electrophoresis . The esterase activity of renal kallikrein determined by apectrophotometric assay, using benzoyl-L-arginine ethyl ester (BAEE) as substrate, was 1650 EU/mg . The esterase unit is arbitrarily defined as that amount of enzyme which hydro lyzes 0.05 uMolea of BAEE/minute at pH 8 .0 and 25 °C . The differences between renal kallikrein and the most abundant rat urinary kallikrein (M .W . 32,000 and 1030 EU/mg, isolated by affinity chromatography from rat urine) suggest that the urinary enzyme is a partial degradation product of renal kallikrein . PIG PANCREATIC KALLIKREIN :
STRUCTURE AND CATALYTIC PROPERTIES F. Fiedler
Institut für Klinische Chemie und Klinische Biochemie der Universität Plünchen, ?tünchen GFR Rallikreins A and B from porcine pancreas have identical amino acid compositions. Both enzymes contain N-terminal Ile and Ala and C-terminal Leu/Ser and Pro. The only difference found so far is roughly double the amount of neutral sugars (mannose, galactose, and fucoae) and of glucosamine and possibly an excess of three amide residues in kallikrein B . In cooperation with Dre . Tschesche, Kutzbach, and Schmidt-Kaetner, we have determined the amino acid sequence of about 1/3 of the kallikrein B molecule . There is evident-a large extent of homology with other pancreatic aerine proteinasea . Kinetic constants for the kallikrein catalyzed hydrolysis of bradykin~l-Ser-Val-Gln and C-terminal
Conference Abstracts
Vol . 16, No . 5
Of the four rat urinary kallikrein fractions noted by Nustad and Pierce (1), the moat abundant was isolated using Whatman DE-32 cellulose and affinity chromatography . The latter involved the use of Aprotinin (TrasylolR) bound by the amide method to Bio-Gel P-200 . Using SDS-polyacrylamide gel electrophoresis, molecular weights of 37,000, 31,000 and 32,000 were estimated respectively for human, rabbit and rat urinary kallikreina . The respective esterase activities were 30, 67, and 1030 esterase unite/mg (apectrophotometric assay using benzoyl-L-arginine ethyl ester) . Conspicuous differences in the amino acid composition of the kallikrein were found. 1.
R. NUSTAD and J .V . PIERCE, Biochemistry Z3 2312-2319 (1974) . PRELIMINARY DATA ON PURIFIED RALLIKREIN FROM RAT KIDNEY G. Porcelli, R.H . Croxatto, G.B . Marini-Hettalo and M . Di Iorio
Instituto di Chimica delle Facoltà di Medicine dell'Univeraità Cattolica, Rome Centro dells Chimica dei Recettori del C .N .R ., Rome Laboratorio de Fiaiologa della Univereidad Catolica, Santiago, Chile A highly purified kallikrein from rat kidney was obtained by a multi-step purification procedure including gel filtration on Sephadex 6-100, chromatography on Whatman CM-32 and DE 32 celluloses and Sephadex G-100 superfine. Molecular weight of 40,000 was estimated for the purified enzyme by sodium dodecyl sulfate polyacrylamide disc gel electrophoresis . The esterase activity of renal kallikrein determined by apectrophotometric assay, using benzoyl-L-arginine ethyl ester (BAEE) as substrate, was 1650 EU/mg . The esterase unit is arbitrarily defined as that amount of enzyme which hydro lyzes 0.05 uMolea of BAEE/minute at pH 8 .0 and 25 °C . The differences between renal kallikrein and the most abundant rat urinary kallikrein (M .W . 32,000 and 1030 EU/mg, isolated by affinity chromatography from rat urine) suggest that the urinary enzyme is a partial degradation product of renal kallikrein . PIG PANCREATIC KALLIKREIN :
STRUCTURE AND CATALYTIC PROPERTIES F. Fiedler
Institut für Klinische Chemie und Klinische Biochemie der Universität Plünchen, ?tünchen GFR Rallikreins A and B from porcine pancreas have identical amino acid compositions. Both enzymes contain N-terminal Ile and Ala and C-terminal Leu/Ser and Pro. The only difference found so far is roughly double the amount of neutral sugars (mannose, galactose, and fucoae) and of glucosamine and possibly an excess of three amide residues in kallikrein B . In cooperation with Dre . Tschesche, Kutzbach, and Schmidt-Kaetner, we have determined the amino acid sequence of about 1/3 of the kallikrein B molecule . There is evident-a large extent of homology with other pancreatic aerine proteinasea . Kinetic constants for the kallikrein catalyzed hydrolysis of bradykin~l-Ser-Val-Gln and C-terminal