PMID:11872174
| Citation |
Kulkarni, GV and Deobagkar, DD (2002) A cytosolic form of aminopeptidase P from Drosophila melanogaster: molecular cloning and characterization. J. Biochem. 131:445-52 |
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| Abstract |
Using a functional genomic approach, we have identified and characterized a cytosolic form of aminopeptidase P from Drosophila melanogaster. This study represents the first characterization of an insect aminopeptidase P. The complete sequence of a 12.5 kbp genomic clone from D. melanogaster showed the presence of a 1,839 bp ORF, encoding a protein of 613 amino acids with a calculated molecular mass of 68.5 kDa. The deduced amino acid sequence was 48% identical and 66% similar to rat and human cytosolic aminopeptidase P. Amino acids important for catalytic activity and the metal binding ligands were found to be conserved between Drosophila AP-P and its mammalian homologues. The recombinant enzyme expressed in Escherichia coli hydrolyzed the amino terminal Xaa-Pro bond of substance P and bradykinin, revealing its functional identity. Further enzyme characterization showed the enzyme to be a manganese-dependent metallopeptidase. Immunoblot analysis showed that DAP-P is located exclusively in the cytosol and is temporally regulated during Drosophila development. In the adult fly, the protein could be detected in gut, testis and ovary, with a high level of expression in brain. |
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| Keywords |
Amino Acid Sequence; Aminopeptidases/genetics; Aminopeptidases/isolation & purification; Aminopeptidases/metabolism; Animals; Base Sequence; Bradykinin/metabolism; Brain/enzymology; Catalytic Domain; Cloning, Molecular; Cytosol/enzymology; DNA/analysis; Drosophila melanogaster/enzymology; Drosophila melanogaster/genetics; Drosophila melanogaster/growth & development; Humans; Manganese/metabolism; Metalloendopeptidases/genetics; Metalloendopeptidases/isolation & purification; Metalloendopeptidases/metabolism; Molecular Sequence Data; Protein Binding; Sequence Homology, Amino Acid; Substance P/metabolism; Substrate Specificity |
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