HALOALKANE DEHALOGENAZES RE-DESIGN
WHERE and WHEN?Loschmidt Laboratories, Masaryk University, Brno, Czech Republic 2010 – 2013
Modification of tunnels in haloalkane dehalogenazes (DatA, LinB) design of new mutants with modified activity and selectivity
Protein engineering can be applied for tailoring of naturally evolved enzymes. Modifications introduced to the residues forming the first or second shell of an active site currently represent the most common strategy for enzyme redesign. Recent project presents an example of engineering of substrate specificity of haloalkane dehalogenase by targeting the residues of their access tunnels.
Protein design was based on computational approach, with combination of QM/MM methods, MD and RAMD simulations, docking and software for tunnels detection (CAVER). Predicted mutants were constructed and their properties were analyzed by several experimental methods.
The haloalkane dehalogenases are the family of enzymes with buried active sites. Their activity and selectivity reflect the properties of the active site as well as substrate/product transport pathways. Project explores possibility of tunnels re-engineering in bacterial enzymes haloalkane dehalogenases. Based on the analysis carried out, new mutants of haloalkane dehalogenases LinB and DatA were designed with improved catalytic properties (activity and selectivity). Work provides evidence that careful redesign of access pathways represents a powerful strategy for the precise control of activity and selectivity of enzymes.
B. Kozlikova, E. Sebestova, V. Sustr, J. Brezovsky, O. Strnad, L. Daniel, D. Bednar, A. Pavelka, M. Manak, M. Bezdeka, P. Benes, M. Kotry, A. Gora, J. Damborsky, and J. Sochor, “CAVER Analyst 1.0: graphic tool for interactive visualization and analysis of tunnels and channels in protein structures,” Bioinforma. Oxf. Engl., vol. 30, no. 18, pp. 2684–2685, Sep. 2014.
A. Gora, J. Brezovsky, and J. Damborsky, “Gates of Enzymes,” Chem. Rev., vol. 113, no. 8, pp. 5871–5923, 2013.
K. Hasan, A. Gora, J. Brezovsky, R. Chaloupkova, H. Moskalikova, A. Fortova, Z. Prokop, J. Damborsky – “Effect of Unique Halide-Stabilising Residue on the Catalytic Properties of Haloalkane Dehalogenase DatA from Agrobacterium tumefaciens C58” – FEBS J., vol. 280, no. 13, pp. 3149–3159, July 2013.
J. Brezovsky, E. Chovancova, A. Gora, A. Pavelka, L. Biedermannova, and J. Damborsky, “Software tools for identification, visualization and analysis of protein tunnels and channels,” Biotechnol. Adv., vol. 31, no. 1, pp. 38–49, Jan. 2013.ARTICLE
E. Chovancova, A. Pavelka, P. Benes, O. Strnad, J. Brezovsky, B. Kozlikova, A. Gora, V. Sustr, M. Klvana, P. Medek, L. Biedermannova, J. Sochor, and J. Damborsky, “CAVER 3.0: A Tool for the Analysis of Transport Pathways in Dynamic Protein Structures,” PLoS Comput Biol, vol. 8, no. 10, p. e1002708, Oct. 2012.ARTICLEOpen Access
L. Biedermannová, Z. Prokop, A. Gora, E. Chovancová, M. Kovács, J. Damborsky, and R. C. Wade, “A single mutation in a tunnel to the active site changes the mechanism and kinetics of product release in haloalkane dehalogenase LinB,” J. Biol. Chem., vol. 287, no. 34, pp. 29062–29074, Aug. 2012.ARTICLE
WHERE and WHEN?
AIST Tohoku – Sendai, JAPAN, 2002/2003 and 2004-2006
Dehydrogenation of metylocyclohexane into toluene
Storage of hydrogen in the form of chemical compounds can be an attractive alternative to traditional methods due to both economic and safety issue. One of the problems that still need to be solved is dehydrogenation of compounds and hydrogen recovery from storage compounds. The project aim development of methodology for easy hydrogen recovery and separation.
By the use of catalysts combined with modern membranes of high durability in low temperatures.
The use of compounds such as toluene (TOL), has been already shown to be potentially feasible for hydrogen storage for both mobile and stationary applications. The reversible reaction of dehydrogenation of MCH to toluene, and in general all dehydrogenation reactions of paraffins to aromatic compounds, is highly endothermic and strongly limited by thermodynamic equilibrium. The ability of the catalyst placed inside membrane tube combined with the selectivity of hydrogen permeation through the membrane makes it possible to remove hydrogen and thereby shift the reaction towards the product side. Careful study allow me to confirm high durability and stability of pore filled type of palladium membrane in reaction conditions in the low temperature range (100-300ºC). The new composition of membrane and location of palladium grains inside pores of ? alumina intermediate layer allowed to use palladium membranes in such drastic conditions without losing their selectivity. So far there was no any other reported information about membrane working in temperature below 300ºC for such a long time.
A. Gora, D. A. P. Tanaka, F. Mizukami, and T. M. Suzuki, “Lower temperature dehydrogenation of methylcyclohexane by membrane-assisted equilibrium shift,” Chem. Lett., vol. 35, pp. 1372–1373, 2006. ARTICLE
WHERE and WHEN?Institute of Catalysis and Surface Chemistry Polish Academy of Sciences and Jagiellonian University – Krakow, POLAND 2003-2005
Study of colorizing and blanching phenomena in “smart windows”
Electrochromic devices can find application in “smart windows” fabrication, windows which are able to decrease the use of electricity consumption for air conditioning. The project aim investigations of the tungsten oxides role in the colorizing and blanching phenomena.
By theoretical methods (quantum chemical calculations – TD-DFT).
Electrochromic devices can find application in “smart windows” fabrication, windows which are able to decrease the use of electricity consumption for air conditioning. The purpose of the research was to investigate the mechanism of WO3 layer colorizing and blanching phenomena. To achieve this, the mechanism of Li ion transfer and its intercalation into WO3 bulk structure was studied by using advanced theoretical tools (TD-DFT methodology).
E. Broclawik, A. Gora, P. Liguzinski, P. Petelenz, and H. A. Witek, “Quantum chemical modeling of electrochromism of tungsten oxide films,” J. Chem. Phys., vol. 124, 2006. ARTICLE
E. Broclawik, A. Gora, P. Liguzinski, P. Petelenz, and M. Slawik, “Quantum chemical modelling of the process of lithium insertion intoWO(3) films,” Catal. Today, vol. 101, pp. 155–162, 2005. ARTICLE
SCR of Nitric Oxides
WHERE and WHEN?Jagiellonian University – Krakow, POLAND 1996-2002
Study of vanadium-tungsten catalysts for DENOX process
Nitric oxides are important air pollutants. Vanadium-tungsten catalysts are commercially used for selective catalytic reduction (SCR) of NOx by ammonia, however the role of tungsten is unclear. The project aim investigations of the tungsten role in the increasing of the catalyst activity in low temperatures.
By the combination of experimental (TEM, Raman spectroscopy, vacuum equipment, GC/MS) and theoretical (quantum chemical calculations – DFT) methods.
Vanadium-tungsten catalysts are commercially used for selective catalytic reduction (SCR) of NOx by ammonia. The role of tungsten in decreasing of the onset temperature of the SCR of NOx remains unclear up to now. This role seems to be essential for better understanding of the catalyzed reaction and future development of more efficient catalysts. To clarify this, I have i) studied the interaction of water molecule with vanadia-tungsten catalyst surface and the creation of new active centers for SCR process by FT Raman spectroscopy and quantum chemical (DFT) modeling ii) successfully synthetized highly active vanadium-tungsten catalysts for SCR of NOx by NH3 by the use of sol-gel method. Extensive calculations for cluster models of active sites on the catalyst surface and their interaction with small substrate molecules showed possibility of the use of water molecules as a probe of acid-base properties of Lewis and Brønsted surface centers.
A. Gora and E. Broclawik, “Theoretical estimation of acid-base properties of Lewis and Bronsted centres at the V-W-O catalyst surface: water molecule as the probe in DFT calculations,” J. Mol. Catal. -Chem., vol. 215, pp. 187–193, 2004. ARTICLE
M. Najbar, A. Gora, A. Bialas, and A. Weselucha-Birczynska, “Low-temperature reactivity of the surface species of vanadia-tungsta catalyst,” Solid State Ion., vol. 141, pp. 499–506, 2001.ARTICLE
E. Broclawik, A. Gora, and M. Najbar, “The role of tungsten in formation of active sites for no SCR on the V-W-O catalyst surface – quantum chemical modeling (DFT),” J. Mol. Catal. -Chem., vol. 166, pp. 31–38, 2001. ARTICLE
M. Najbar, E. Broclawik, A. Gora, J. Camra, A. Bialas, and A. Weselucha-Birczynska, “Evolution of the surface species of the V2O5-WO3 catalysts,” Chem. Phys. Lett., vol. 325, pp. 330–339, 2000. ARTICLE
A. Gora, E. Broclawik, “Dissociation of the Water Molecule on the V-W-O Catalyst Surface – Quantum Chemical Modeling”, Polish Journal of Environmental Studies, 2000; 9(1):31-34.
A. Gora, E. Broclawik, and M. Najbar, “Quantum chemical modeling (DFT) of active species on the V-W-O catalyst surface in various redox conditions,” Comput. Chem., vol. 24, pp. 405–410, 2000. ARTICLE
M. Najbar, F. Mizukami, A. Bialas, J. Camra, A. Weselucha-Birczynska, H. Izutsu, and A. Gora, “Evolution of Ti-Sn-rutile-supported V2O5-WO3 catalyst during its use in nitric oxide reduction by ammonia,” Top. Catal., vol. 11, pp. 131–138, 2000. ARTICLE
M. Najbar, A. Bialas, F. Mizukami, A. Weselucha-Birczynska, E. Bielanska, A. Gora, “Vanadia-Tungsta DENOX Catalysts on High Surface Area Rutile”, Polish Journal of Environmental Studies, 1997; 6:83-88.