This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. NAD(P)H-dependent oxidoreductases are valuable tools for synthesis of chiral compounds. The expensive cost of the cofactors, however, requires in situ cofactor regeneration for preparitive applications. We have attempted to develop an enzymatic system based on phosphite dehydrogenase from Pseudomonas stutzeri to regenerate the reduced nicotinamide cofactors NADH and NADPH. Directed evolution has been used to generate mutants with higher thermal stability. Techniques such as Circular Dichroism and Differential Scanning Calorimetry will be used to elucidate the mechanism of individual stabilizing mutations.
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