CD Spectroscopy of Nucleic Acids and Proteins
Prof. RNDr. Michaela Vorlíčková, DrSc.
THEMATIC RESEARCH FOCUS
- Studies in the conformational properties of genomic DNA fragments important from biological and medical points of view
- Investigations into the structures and interactions of biomacromolecules and how they relate to the functions of living systems, diseases and therapies
- To map the conformational properties of DNA sequence motifs occurring in important regions of the human genome.
- To characterize the phyico-chemical properties of non-B DNA structures, chemical alterations in DNA, and other external factors affecting their stability, dynamics and recognition by proteins.
CONTENT OF RESEARCH
Depending on sequence, DNA can adopt various secondary structures distinct from Watson and Crick’s classic double helix. The conformational distinctions include double helix handedness, orientation of strands, type of base pairing, and the number of associated strands. These unusual secondary structures are frequently implicated in various biological functions or diseases. At present, the most frequently studied unusual secondary structures are quadruplexes.
There are numerous regions in the human genome prone to forming quadruplexes. They frequently occur in gene promoters and control their functions. We focus on quadruplexes formed by telomeric DNA and DNA of selected oncogenes. The quadruplexes formed in telomeres influence aging and cancer, and thus have become targets for the design of anticancer agents.
A further ‘hot topic’ is currently the reprogramming of differentiated cells into pluripotent stem cells. The most important pluripotency determinants, c-myc and oct4 genes, contain a DNA sequence in their promotor regions that is capable of forming quadruplex. Our expertise in the prediction of quadruplex formation, in their experimental detection and characterization in relation to their function will become the basis for perspective development of targeted therapy for cancer and other diseases.
The main method used by the group is electronic circular dichroism (CD) spectroscopy. It is extremely sensitive to changes in mutual orientation of DNA constituents and, therefore, it is uniquely suited for DNA conformational studies. Particular DNA arrangements (classical B- and A-forms, left-handed Z-form, parallel and antiparallel guanine quadruplexes or intercalated cytosine tetraplexes) provide characteristic CD spectra.
KEY RESEARCH EQUIPMENT
CURRENT RESEARCH INFRASTRUCTURE
The research group is equipped with UV/VIS absorption spectrophotometer Varian Cary 4000; Jobin-Yvon Mark VI and Jasco 185 CD spectrometers; Temperature gradient gel electrophoresis Model TGGE Maxi system.
- DNA quadruplexes in human genome associated with disease and aging (GAP205/12/0466) Czech Science Foundation, 2012-2016, M. Vorlickova, Institute of Biophysics AS CR.
- Biophysics and bioinformatics of genomic DNA fragments very rich in guanine and adenine (IAA500040903), Academy of Sciences of the Czech Republic, 2009-2013, J. Kypr, Institute of Biophysics AS CR, V. Sklenar, Masaryk University.
- Formation and dynamics of nucleic acid motifs involved in regulation of gene expression (GA202/09/0193), Czech Science Foundation, 2009-2014, J. Stepanek, Charles University in Prague.
- VORLICKOVA, M., KEJNOVSKA, I., SAGI, J., RENCIUK, D., BEDNAROVA. K., MOTLOVA, J., KYPR, J. Circular dichroism and guanine quadruplexes. Methods. 2012, 57, p. 64–75.
- KYPR, J., KEJNOVSKA, I., BEDNAROVA, K., VORLICKOVA, M. Circular dichroism spectroscopy of nucleic acids. In Comprehensive chiroptical spectroscopy: Application in stereochemical analysis and synthetic compounds, natural products, and biomolecules. N. Berova, P.L. Polavarapu, K. Nakanishi, R.W. Woody. Eds. John Wiley & Sons. Inc., 2012. p. 575-586.
- KYPR, J., KEJNOVSKA, I., RENCIUK, D., VORLICKOVA, M. Circular dichroism and conformational polymorphism of DNA. Nucleic Acids Research. 2009, 37(6), p. 1713-1725.
- RENCIUK, D., KEJNOVSKA, I., SKOLAKOVA, P., BEDNAROVA, K., MOTLOVA, J., VORLICKOVA, M. Arrangements of human telomere DNA quadruplex in physiologically relevant K+ solutions. Nucleic Acids Research. 2009, 37(19), p. 6625-6634.
- VORLICKOVA, M., CHLADKOVA, J., KEJNOVSKA, I., FIALOVA, M., KYPR, J. Guanine tetraplex topology of human telomere DNA is governed by the number of (TTAGGG) repeats. Nucleic Acids Research. 2005, 33(18), p. 5851-5860.