Description of Transport Tunnel in Haloalkane Dehalogenase Variant LinB D147C+L177C from Sphingobium japonicum

Authors

Iermak, I., Degtjarik, O., Havlickova, P., Kuty, M., Chaloupkova, R., Damborsky, J., Prudnikova, T., Kuta Smatanova, I.

Source

CATALYSTS 11: 5 (2021)

Abstract

Activity of enzymes with active sites buried inside the protein core highly depends on the efficient transport of substrates and products between the active site and the bulk solvent. Engineering of access tunnels in order to increase or decrease catalytic activity and specificity in a rational way is a challenging task. Here we describe a combined experimental and computational approach to characterize the structural basis of altered activity in the haloalkane dehalogenase LinB D147C+L177C variant. While the overall protein fold is similar to the wild type enzyme and the other LinB variants, the access tunnels have been altered by introduced cysteines that were expected to form a disulfide bond. Surprisingly, the mutations have allowed several conformations of the amino acid chain in their vicinity, interfering with the structural analysis of the mutant by X-ray crystallography. Time required for growing of protein crystals changed from days to 1.5 years by introducing the substitutions. The haloalkane dehalogenase LinB D147C+L177C variant crystal structure was solved to 1.15 Å resolution, characterized and deposited to Protein Data Bank under PDB ID 6S06. 

Full text

Citation

Iermak, I., Degtjarik, O., Havlickova, P., Kuty, M., Chaloupkova, R., Damborsky, J., Prudnikova, T., Kuta Smatanova, I., 2021: Description of Transport Tunnel in Haloalkane Dehalogenase Variant LinB D147C+L177C from Sphingobium japonicum. Catalysts 11: 5.

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