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Article

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

1
Department of Structural Cell Biology, Max Planck Institute of Biochemistry, Am Klopferspitz 18, 82152 Martinsried, Germany
2
Department of Structural Biology, Weizmann Institute of Science, Rehovot 7610001, Israel
3
Faculty of Science, University of South Bohemia in Ceske Budejovice, Branisovska 1760, 37005 Ceske Budejovice, Czech Republic
4
Loschmidt Laboratories, Department of Experimental Biology and RECETOX, Faculty of Science, Masaryk University, Kamenice 5, 62500 Brno, Czech Republic
5
International Clinical Research Center, St. Anne’s University Hospital Brno, Pekarska 53, 65691 Brno, Czech Republic
*
Authors to whom correspondence should be addressed.
Catalysts 2021, 11(1), 5; https://doi.org/10.3390/catal11010005
Received: 20 November 2020 / Revised: 13 December 2020 / Accepted: 18 December 2020 / Published: 23 December 2020
(This article belongs to the Special Issue Catalysts for Structure-Functional Analysis and Enzyme Optimization)
The activity of enzymes with active sites buried inside their protein core highly depends on the efficient transport of substrates and products between the active site and the bulk solvent. The 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. The duration required for the 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. View Full-Text
Keywords: bacterial enzyme; haloalkane dehalogenase; mutant form; crystallization; tertiary structure; disulfide bond; protein engineering; molecular dynamics; access tunnel; substrate specificity bacterial enzyme; haloalkane dehalogenase; mutant form; crystallization; tertiary structure; disulfide bond; protein engineering; molecular dynamics; access tunnel; substrate specificity
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MDPI and ACS Style

Iermak, I.; Degtjarik, O.; Havlickova, P.; Kuty, M.; Chaloupkova, R.; Damborsky, J.; Prudnikova, T.; Kuta Smatanova, I. Description of Transport Tunnel in Haloalkane Dehalogenase Variant LinB D147C+L177C from Sphingobium japonicum. Catalysts 2021, 11, 5. https://doi.org/10.3390/catal11010005

AMA Style

Iermak I, Degtjarik O, Havlickova P, Kuty M, Chaloupkova R, Damborsky J, Prudnikova T, Kuta Smatanova I. Description of Transport Tunnel in Haloalkane Dehalogenase Variant LinB D147C+L177C from Sphingobium japonicum. Catalysts. 2021; 11(1):5. https://doi.org/10.3390/catal11010005

Chicago/Turabian Style

Iermak, Iuliia, Oksana Degtjarik, Petra Havlickova, Michal Kuty, Radka Chaloupkova, Jiri Damborsky, Tatyana Prudnikova, and Ivana Kuta Smatanova. 2021. "Description of Transport Tunnel in Haloalkane Dehalogenase Variant LinB D147C+L177C from Sphingobium japonicum" Catalysts 11, no. 1: 5. https://doi.org/10.3390/catal11010005

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