Balancing the Stability-Activity Trade-off by Fine-Tuning Dehalogenase Access Tunnels

Authors

Lišková, V., Bednář, D., Holubeva, T., Prudniková, T., Řezáčová, P., Koudeláková, T., Šebestová, E., Kutá Smatanová, I., Brezovský, J., Chaloupková, R., Damborský, J.

Source

ChemCatChem 7(4)

Abstract

A variant of the haloalkane dehalogenase DhaA with greatly enhanced stability and tolerance of organic solvents but reduced activity was created by mutating four residues in the access tunnel. To create a stabilised enzyme with superior catalytic activity, two of the four originally modified residues were randomised. The resulting mutant F 176 G exhibited 32- and 10-times enhanced activity towards 1,2-dibromoethane in buffer and 40 % DMSO, respectively, upon retaining high stability. Structural and molecular dynamics analyses demonstrated that the new variant exhibited superior activity because the F 176 G mutation increased the radius of the tunnel’s mouth and the mobility of α-helices lining the tunnel. The new variant’s tunnel was open in 48 % of trajectories, compared to 58 % for the wild-type, but only 0.02 % for the original four-point variant. Delicate balance between activity and stability of enzymes can be manipulated by fine-tuning the diameter and dynamics of their access tunnels.

Source

Lišková, V., Bednář, D., Holubeva, T., Prudniková, T., Řezáčová, P., Koudeláková, T., Šebestová, E., Kutá Smatanová, I., Brezovský, J., Chaloupková, R., Damborský, J.: Balancing the Stability-Activity Trade-off by Fine-Tuning Dehalogenase Access Tunnels, ChemCatChem, 7(4), 648–659, 2015.
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