Using singular perturbation theory to determine kinetic parameters in a non-standard coupled enzyme assay

• DOI: 10.1007/s00285-020-01524-8
• Publication Date: 2020-08-06T11:02:55Z
• Authors: Mohit P. Dalwadi; Diego Orol; Frederik Walter; Nigel P. Minton; John R. King; Katalin Kovács
• Keywords: Cupriavidus necator

Abstract We investigate how to characterize the kinetic parameters of an aminotransaminase using a non-standard coupled (or auxiliary) enzyme assay, where peculiarity arises for two reasons. First, one products auxiliary is substrate primary and, second, we explicitly account reversibility reaction. Using singular perturbation theory, distinguished asymptotic limits in terms strength reverse reaction, which allows us determine deduce characterized enzyme. This establishes parameter-estimation algorithm that applicable more generally similar reaction networks. demonstrate applicability our theory by performing assays novel putative enzyme, CnAptA (UniProtKB Q0KEZ8) from Cupriavidus necator H16, different omega-amino acid substrates.