title: An analytical method for determining relative specificities for sequential reactions catalyzed by the same enzyme: general formulation.
abstract: We present a general formulation of a model that can be used to analyze reaction profiles in systems in which a single enzyme catalyzes several sequential reactions with the same molecular backbone. The analysis of these so-called "repeated-attack systems" allows estimation of the specificities that the enzyme has for the various intermediate substrates that appear in the reaction mixture, relative to the specificity that it has for the initial substrate. Our analytical method has the important advantage that it is not affected by competitive or uncompetitive inhibition, nor by denaturation of the enzyme during the reaction. We carry out case studies in three different systems, the lipase-catalyzed alcoholysis of triacylglycerols, the phytase-catalyzed removal of phosphate groups from phytic acid and the beta-amylase-catalyzed removal of maltose units from maltoheptaose. Our model fits well to all reaction profiles in which the phenomenon of processivity does not occur. It can therefore be used as a general tool for characterizing the relative specificities of "repeated-attack enzymes".
authors.affiliation: Departamento de Bioquímica e Biologia Molecular, Universidade Federal do Paraná, Cx P 19046 Centro Politécnico, Curitiba 81531-990, Paraná, Brazil. davidmitchell@ufpr.br
authors.forename: David Alexander
authors.initials: DA
authors.lastname: Mitchell
authors.forename: Frédéric
authors.initials: F
authors.lastname: Carrière
authors.forename: Nadia
authors.initials: N
authors.lastname: Krieger
mesh_terms: D000465:Algorithms; D002384:Catalysis; D004798:Enzymes / Q000378:metabolism*; D006868:Hydrolysis; D007700:Kinetics; D008954:Models, Biological; D013379:Substrate Specificity
chemical_list: D004798:Enzymes