tailieunhanh - Báo cáo khoa học: Characterization of a membrane-bound aminopeptidase purified from Acyrthosiphon pisum midgut cells A major binding site for toxic mannose lectins

A single membrane-bound aminopeptidase N (APN) occurs in the pea aphid (Acyrthosiphon pisumHarris) midgut, with a pH optimum of , pI of and molecular mass of 130 kDa. This enzyme accounts for more than of the total gut proteins. After being solubilized in detergent, APN was purified to homogeneity. The enzyme is a glycoprotein rich in mannose residues, which binds the entomotoxic lectins of the concanavalin family. | ễFEBS Journal Characterization of a membrane-bound aminopeptidase purified from Acyrthosiphon pisum midgut cells A major binding site for toxic mannose lectins Plinio T. Cristofoletti1 Flavia A. Mendonọa de Sousa2 Yvan Rahbé2 and Walter R. Terra1 1 Departamento de Bioquimica Institute de Quimica Universidade de Sao Paulo Brazil 2 UMR INRA-INSAde Lyon BF2I Biologie Fonctionnelle Insectes Interactions Villeurbanne France Keywords aminopeptidase N Aphididae glycosyl-phosphatidylinositol GPI anchor mannose lectin receptor substrate specificity Correspondence Y. Rahbe UMR INRA-INSA de Lyon BF2I Biologie Fonctionnelle Insectes Interactions Bat. Louis-Pasteur F-69621 Villeurbanne cedex France Fax 33 4 72 43 85 34 Tel 33 4 72 43 83 56 E-mail Database The sequence described here has been deposited in the GenBank database with the accession number DQ440823 Received 25 August 2006 revised 13 October 2006 accepted 19 October 2006 doi A single membrane-bound aminopeptidase N APN occurs in the pea aphid Acyrthosiphon pisum Harris midgut with a pH optimum of pI of and molecular mass of 130 kDa. This enzyme accounts for more than of the total gut proteins. After being solubilized in detergent APN was purified to homogeneity. The enzyme is a glycoprotein rich in mannose residues which binds the entomotoxic lectins of the concanavalin family. The internal sequence of APN is homologous with a conservative domain in APNs and degenerated primers of highly conserved APN motifs were used to screen a gut cDNA library. The complete sequence of APN has standard residues involved in zinc co-ordination and catalysis and a glycosyl-phosphatidylinositol anchor as in APNs from Lepidoptera. APN has a broad specificity towards N-terminal amino acids but does not hydrolyze acidic aminoacyl-peptides thus resembling the mammalian enzyme EC . The kcat Km ratios for different di- tri- tetra- and penta-peptides suggest a .