PURIFICATION AND PROPERTIES OF DIPEPTIDYL AMINOPEPTIDASE-I FROM CHICKEN LIVER

  • 1 January 1980
    • journal article
    • research article
    • Vol. 36  (3) , 321-330
Abstract
Dipeptidyl aminopeptidase I (E.S. 3.4.14.1) from chicken liver was purified by the following steps: homogenization at pH 5, thermic precipitation, acetone fractionation and Sephadex G-100, DEAE-cellulose and organomercurial-Sepharose column fractionations. The purified enzyme appears to be homogeneous by polyacrylamide gel electrophoresis at both pH 4.5 and 8.3 and has an isoelectric point of 5.7 .+-. 0.05. The MW of the enzyme reale 167,000 .+-. 17,000 on the Sephadex G-150 column chromatography. The optimum pH for hydrolysis of Gly-Phe-p-nitroanilide (GPNA) and Gly-Phe-B-naphthylamide was 5.8. The value of Km for the hydrolysis of GPNA was estimated at 3.3 mM. The enzyme required halide ions for activity and was activated by thiol reagents (dithiothreitol, cysteine and 2-mercaptoethanol). DAP I was inhibited by thiol-blocking reagents (PCMB, IAA and Hg2+). The enzyme oxidation with oxygen current was fostered by chloride anion (50 mM); the activity was recovered when cysteine was present in the incubation mixture; the latter seems to perform as enzyme protector.

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