Purification and partial characterization of cathepsin D from porcine (Sus scrofa) liver using affinity chromatography

Cathepsin D, a lysosomal aspartic protease, has been purified from porcine liver using a combination of pepstatin‐A agarose and Affi‐Gel Blue affinity chromatography, followed by size‐exelusion chromatography. The purified protein consists of two polypeptide chains of 15 and 30 kDa, and has an isoel...

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Bibliographic Details
Published in:Biochemistry and molecular biology international 1998-07, Vol.45 (4), p.797-803
Main Authors: Canduri, Femanda, Ward, Richard J., de Azevedo, Walter F., Gomes, Roseli A. S., Arni, R. K.
Format: Article
Language:English
Subjects:
affinity chromatography
Animals
aspartic protease
Cathepsin D
Cathepsin D - chemistry
Cathepsin D - isolation & purification
Cathepsin D - metabolism
Chromatography, Affinity
Electrophoresis, Polyacrylamide Gel
Hemoglobins - metabolism
Hydrogen-Ion Concentration
Isoelectric Point
Kinetics
Liver - enzymology
lysosomal enzyme
Molecular Weight
pepstatin A
Pepstatins
Peptides - metabolism
Swine
Triazines
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Summary:Cathepsin D, a lysosomal aspartic protease, has been purified from porcine liver using a combination of pepstatin‐A agarose and Affi‐Gel Blue affinity chromatography, followed by size‐exelusion chromatography. The purified protein consists of two polypeptide chains of 15 and 30 kDa, and has an isoelectric point of 6.8. Porcine fiver cathepsin D has maximum activity at pH 2.5‐3.0 as determined by its activity against hemoglobin, with a Kcat of 14.3 s‐1 and a kcat/KM of 2.70×106 s‐1M‐1 as determined by the hydrolysis of a fluorogenic peptide substrate.
ISSN:1521-6543
1039-9712
1521-6551
DOI:10.1080/15216549800203222