Mechanistic model for the synthesis of N-acetylneuraminic acid using N-acetylneuraminate lyase from Escherichia coli K12

N-Acetylneuraminate lyase (NAL) from Escherichia coli K12 is an important enzyme for the production of N-acetylneuraminic acid (Neu5Ac), catalyzing the reversible aldol condensation between N-acetyl-D-mannosamine (ManNAc) and pyruvate. Despite the industrial importance of this enzyme, its kinetic mechanism has never been elucidated before. The initial rate patterns were consistent with a rapid-equilibrium ordered bi uni mechanism with pyruvate binding first. Based on progress curve analysis, a mechanistic model was developed to predict the reaction course of Neu5Ac synthesis. The model accurately reproduced the experimental data in a wide range of initial conditions. The correct assignment of the kinetic mechanism is a critical element in optimizing enzymatic syntheses by means of mathematical models, which have become indispensable tools for the design of cost-effective biocatalytic processes. (C) 2012 Published by Elsevier B.V.