Expression, purification, and characterization of an aminopeptidase (Xac2987) with broad specificity from Xanthomonas axonopodis pv. citri.
We report here, the cloning, expression, and purification of a broad specificity aminopeptidase from Xanthomonas campestris pv. citri in fusion with a hexa-histidine tag at the N-terminal portion of the protein to facilitate purification. The protein was expressed in the soluble fraction and could be purified in one step by IMAC, yielding approximately 50mg pure protein per liter of cells. We show that the protein is folded and presents aminopeptidase activity against synthetic substrates. Also, we present the characterization of its specificity, showing that the protein was, indeed, able to catalyze the removal of N-terminal residues from synthetic substrates.
Santos
Kelly
K
Centro de Biologia Molecular Estrutural, Laboratório Nacional de Luz Síncrotron, Caixa Postal 6192, CEP 13084-971 Campinas, Brazil.
Medrano
Francisco J
FJ
D000595:Amino Acid Sequence; D000626:Aminopeptidases / Q000737:chemistry / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism*; D001483:Base Sequence; D017124:Conserved Sequence; D017931:DNA Primers; D007700:Kinetics; D008969:Molecular Sequence Data; D011994:Recombinant Proteins / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism; D016415:Sequence Alignment; D017385:Sequence Homology; D013056:Spectrophotometry, Ultraviolet; D013379:Substrate Specificity; D053532:Xanthomonas axonopodis / Q000201:enzymology* / Q000235:genetics
D017931:DNA Primers; D011994:Recombinant Proteins; D000626:Aminopeptidases