Purification and Characterization of an Aminopeptidase from Lactococcus lactis subsp. cremoris AM2
AUTOR(ES)
Neviani, E.
RESUMO
An aminopeptidase was purified from cell extracts of Lactococcus lactis subsp. cremoris AM2 by ion-exchange chromatography. After electrophoresis of the purified enzyme in the presence or absence of sodium dodecyl sulfate, one protein band was detected. The enzyme was a 300-kilodalton hexamer composed of identical subunits not linked by disulfide bridges. Activity was optimal at 40°C and pH 7 and was inhibited by classical thiol group inhibitors. The aminopeptidase hydrolyzed naphthylamide-substituted amino acids, as well as dipeptides and tripeptides. Longer protein chains such as the B chain of insulin were hydrolyzed, but at a much slower rate. The Michaelis constant (Km) and the maximal rate of hydrolysis (Vmax) were, respectively, 4.5 mM and 3,600 pkat/mg for the substrate l-histidyl-β-naphthylamide. Amino acid analysis showed that the enzyme contained low levels of hydrophobic residues. The partial N-terminal sequence of the first 19 residues of the mature enzyme was determined. Polyclonal antibodies were obtained from the purified enzyme, and after immunoblotting, there was no cross-reaction between these antibodies and other proteins in the crude extract.
ACESSO AO ARTIGO
http://www.pubmedcentral.nih.gov/articlerender.fcgi?artid=203073Documentos Relacionados
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