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X-ray Crystallography I

Doc. RNDr. Jaromír Marek, Ph.D.
Research Group Leader

THEMATIC RESEARCH FOCUS

RESEARCH AREAS

  • Crystallisation of biological macromolecules
  • Determination of 3D structures of molecules, biological macromolecules and macromolecular assemblies by single-crystal diff raction of X-ray
  • Determination of the shape and size of biological macromolecules by small angle X-ray scattering (SAXS)

MAIN OBJECTIVES

  • High throughput structural characterisation of molecular and macromolecular assemblies by single-crystal X-ray diff raction or by SAXS.
  • Investigations of the structure and interactions of biomacromolecules and their relation to the functions of living systems, disease and therapy.

CONTENT OF RESEARCH

The diffraction of X-ray is the main experimental technique used for determination of 3D structures of proteins and their complexes to atomic and subatomic resolution.

The principal bottleneck of the method – the necessity for single crystals of the studied compounds – is now more or less overcome by the employment of highly automated high throughput robotised instrumentation capable of testing simultaneously hundreds and thousands of diff erent crystallisation conditions. The same instrumentation also signifi cantly improves the speed (or the lack of it) of crystallisation screening by changing the scale of crystallisation trials from micro-litre volumes used by human experimenters to nano-litre scale available only to robotised pipettors.

The relative deficiency of experimental time for diff raction experiments is now addressed by using automation for diff raction experiments. Automated diff ractometers can rapidly test the diff raction quality of many available crystals and select the best candidates among them for complete, time-consuming diffraction experiments either at an in-house diff ractometer or at a distant synchrotron radiation facility.

Regardless the automation of hardware and improvement of software abilities, methods for obtaining of 3D structures by single crystal diffraction of X-ray (or by analysis of NMR data) fail quite often. The time and money spent on protein sample preparation are usually wasted in such cases. Fortunately, there is another alternate technique capable of obtaining at least a low resolution 3D shape of the macromolecule/macromolecular complex even from problematic liquid samples: small angle X-ray scattering (SAXS).

The new equipment planned for the CEITEC Single Crystal X-ray Diffraction Core Facility will allow us to use all the above described approaches and implement X-ray crystallography and SAXS as widely-used methodologies for the study of (bio)chemical and biological processes at the atomic or nearly atomic level available for the majority of CEITEC research programmes.

KEY RESEARCH EQUIPMENT

PLANNED RESEARCH INFRASTRUCTURE

Core Facility

The research group will be one of the principal users of the CEITEC Single Crystal X-ray Diffraction Core Facility.

CURRENT RESEARCH INFRASTRUCTURE

The existing infrastructure of the research group of single crystal X-ray diffraction consists of a 4-circle diffractometer (Kuma/Oxford Diffraction/Varian/Agilent) working with a conventional source of X-ray (a sealed tube, mainly with „inorganic“ wavelength of Mo Kα), a CCD area detector and low temperature equipment (Oxford Cryosystems) for work at cryotempatures.

MAIN PROJECTS

  • Synthesis and properties of inorganic-organic polymeric materials (GA203/08/1111), Czech Science Foundation, 2008-2010, M. Nečas, Masaryk University.
  • Controlled synthesis of glycoluril supramolecular objects (GAP207/10/0695), Czech Science Foundation, 2010-2013, V. Šindelař, Masaryk University.
  • Lectins from human pathogens – structure, function, engineering (GA303/09/1168), Czech Science Foundation, 2009-2012, M. Wimmerova, Masaryk University.
  • Design of Carbohydrates and Glycomimetics as Antibacterial and Antiviral Drugs (ME08008), Ministry of Education, Youth and Sports, 2008-2012, M. Wimmerova, Masaryk University.
  • Structure-functional characterization of oxidoreductases acting on nitrogenous regulatory compounds in plants (GA522/08/0555), Czech Science Foundation, 2008-2012, M. Šebela, Palacky University Olomouc, M. Wimmerova, Masaryk University.

SELECTED PUBLICATIONS

  • PEKAROVA, B., KLUMPLER, T., TRISKOVA, O., HORAK, J., JANSEN, S., DOPITOVA, R., PAPOUSKOVA, V., NEJEDLA, E., ZIDEK, L., SKLENAR, V., MAREK, J., HEJATKO, J., JANDA, L. Structure and binding specificity of the receiver domain of sensor histidine kinase CKI1 from Arabidopsis thaliana. Plant J. 2011, 67, 827-839.
  • HUDSON, J., J., R., BEDNAROVA, K., KOZAKOVA, L., LIAO, C., Y., GUERINEAU, M., COLNAGHI, R., VIDOT, S., MAREK, J., BATHULA, S., R., LEHMANN, A., R., PALECEK, J. Interactions between the Nse3 and Nse4 Components of the SMC5-6 Complex Identify Evolutionarily Conserved Interactions between MAGE and EID Families. PLOS ONE 6. 2011, 6(2), p. e17270.
  • SVEC, J., NECAS, M., SINDELAR, V. Bambus[6]uril. Angewandte Chemie-International Edition. 2010, 49(13), p. 2378-2381.
  • JERABKOVA, B., MAREK, J., BUCKOVA, H., KOPECKOVA, L., VESELY, K., VALICKOVA, J., FAJKUS, J., FAJKUSOVA, L. Keratin mutations in patients with epidermolysis bullosa simplex: correlations between phenotype severity and disturbance of intermediate filament molecular structure. Br. J. Dermatol. 2010, 162(5), p. 1004-1013.
  • KOLMAN, V., MAREK, R., STRELCOVA, Z., KULHANEK, P., NECAS, M., SVEC, J., SINDELAR, V. Electron Density Shift in Imidazolium Derivatives upon Complexation with Cucurbit[6]uril. Chem. Eur. J. 2009, 15(28), p. 6926-6931.
 

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