Rat intestinal brush border membrane dipeptidyl-aminopeptidase IV: kinetic properties and substrate specificities of the purified enzyme |
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Authors: | A M Bella R H Erickson Y S Kim |
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Institution: | 1. Gastrointestinal Research Laboratory (151M2), Veterans Administration Medical Center, 4150 Clement Street, San Francisco, California 94121 U.S.A.;2. University of California, Department of Medicine, San Francisco, California 94143 U.S.A. |
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Abstract: | The kinetic properties and substrate specificities of dipeptidyl-aminopeptidase IV (EC 3.4.14.—) detergent-solubilized and purified from the brush border membrane of rat small intestinal mucosal cells were investigated. Kinetic analysis of purified dipeptidyl-aminopeptidase IV was carried out with a variety of oligopeptides and β-napthylamide derivatives as substrates. In general, peptides with proline penultimate to the amino terminus (XPro, X= amino acid) are more favored substrates while those with alanine (XAla) are hydrolyzed at a slower rate. There is some activity toward substrates having leucine at both the penultimate position and amino terminus (LeuLeu). The activity of the purified enzyme toward GlylProβ-napthylamide derivative is maximal at pH 8.4 in Tris-HCl buffer, with an activation energy of 7.98 kcal/mol. There is no requirement for metal ion. The ability of various dipeptides to inhibit Gly-l-Pro-β-napthylamide derivative hydrolysis was used to determine the binding specificity of the enzyme for the amino-terminal amino acid. These data show that a free amino acid group is necessary for enzymatic activity and increased hydrophobicity of the amino acid at the amino terminus enhances binding. |
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Keywords: | To whom all correspondence should be addressed: Gastrointestinal Research Laboratory |
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