Molecular dynamics provides insights into an engineered oxidoreductase with altered cofactor specificity

Conference Dates

September 15-19, 2019


The aldehyde dehydrogenase from Thermoplasma acidophilum is one of the key enzymes in a previously established synthetic cell-free reaction cascade for the production of alcohols. In order alter the cofactor specificity of this enzyme from NADP+ to NAD+, we applied the CSR-salad tool and investigated further amino acid positions based on its crystal structure. Introduction of five point mutations reduced the Km for NAD+ from 18 mM to 0.6 mM and simultaneously increased the activity for D-glyceraldehyde from 0.4 U/mg to 1.5 U/mg. In order to understand the structural basis of the beneficial mutations, we performed molecular dynamics simulations that showed a significant flexibility gain at the cofactor binding site of the final variant. This increased flexibility facilitates a loop movement that largely contributes to the gain in activity and cofactor specificity. We envision a future optimization potential for aldehyde dehydrogenases based on our results.

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