Mechanistic kinetic model for symmetric carboligations using benzaldehyde lyase.
For reactions using thiamine diphosphate (ThDP)-dependent enzymes many empirically-derived kinetic models exist. However, there is a lack of mechanistic kinetic models. This is especially true for the synthesis of symmetric 2-hydroxy ketones from two identical aldehydes, with one substrate acting as the donor and the other as the acceptor. In this contribution, a systematic approach for deriving such a kinetic model for thiamine diphosphate (ThDP)-dependent enzymes is presented. The derived mechanistic kinetic model takes this donor-acceptor principle into account by containing two K(m)-values even for identical substrate molecules. As example the stereoselective carbon-carbon coupling of two 3,5-dimethoxy-benzaldehyde molecules to (R)-3,3',5,5'-tetramethoxy-benzoin using benzaldehyde lyase (EC 4.1.2.38) from Pseudomonas fluorescens is studied. The model is derived using a model-based experimental analysis method which includes parameter estimation, model analysis, optimal experimental design, in silico experiments, sensitivity analysis and model revision. It is shown that this approach leads to a robust kinetic model with accurate parameter estimates and an excellent prediction capability.
Zavrel
Michael
M
Biochemical Engineering, RWTH Aachen University, Worringerweg 1, D-52074 Aachen, Germany. michael.zavrel@avt.rwth-aachen.de
Schmidt
Thomas
T
Michalik
Claas
C
Ansorge-Schumacher
Marion
M
Marquardt
Wolfgang
W
Büchs
Jochen
J
Spiess
Antje C
AC
D017766:Acid Anhydride Hydrolases / Q000737:chemistry*; D000446:Aldehyde-Lyases / Q000737:chemistry*; D002244:Carbon / Q000737:chemistry*; D003198:Computer Simulation; D007700:Kinetics; D008956:Models, Chemical; D011551:Pseudomonas fluorescens / Q000201:enzymology*
D002244:Carbon; D017766:Acid Anhydride Hydrolases; C023837:nucleoside-diphosphatase; D000446:Aldehyde-Lyases; C059416:benzaldehyde lyase