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Micellar electrokinetic chromatography for analyzing active site specificity of Pseudomonas aeruginosa elastase
Author(s) -
Viglio Simona,
Zanaboni Giuseppe,
Lupi Anna,
Gianelli Luca,
Luisetti Maurizio,
Casali Lucio,
Cetta Giuseppe,
Iadarola Paolo
Publication year - 1999
Publication title -
electrophoresis
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.666
H-Index - 158
eISSN - 1522-2683
pISSN - 0173-0835
DOI - 10.1002/(sici)1522-2683(19990601)20:7<1578::aid-elps1578>3.0.co;2-u
Subject(s) - micellar electrokinetic chromatography , enzyme kinetics , chemistry , elastase , substrate (aquarium) , kinetics , chromatography , cleavage (geology) , catalysis , peptide , active site , pseudomonas aeruginosa , stereochemistry , amino acid , enzyme , electrophoresis , biochemistry , bacteria , materials science , biology , physics , quantum mechanics , ecology , genetics , fracture (geology) , composite material
The geometry of the catalytic site of Pseudomonas aeruginosa elastase was reexamined, exploiting the specific feature of micellar electrokinetic chromatography (MEKC), i.e. , its ability to detect a decrease of intact substrate and simultaneous formation of reaction products. We carried out a detailed investigation using two tri‐ and six tetra‐peptide 4‐nitroanilides (NA) differing from each other by only one or more amino acids as stable substrates. The kinetic cleavage parameters K m and k cat determined by MEKC and the catalytic efficiency K m / k cat values calculated allowed us to better define the substrate specificity of this proteinase.