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Study of structural and binding properties of TRPM channels
Grant agency: GACR
Identification number of grant: 17-04236S
Head of the project: Ing. Jan Teisinger, CSc (Fyziologický ústav AV ČR, v. v. i.)
Vice-head of the project:
RNDr. Vladimír Kopecký Jr., PhD.
Grant annotation
TRP channels are cellular sensors for a wide spectrum of physical and chemical stimuli. Phosphatidylinositol–4,5-bisphosphate (PIP2) is a intracellular messenger which directly interacts with many different effector proteins. The enormous diversity of PIP2 effector proteins allow PIP2 signaling to control a broad range of cellular functions. TRP channels gating processes are regulated by PIP2 binding. The aims of this project will be identification binding sites for PIP2 in both intracellular N- and C-termini of TRPM channels. Fusion proteins of different lengths and mutations in N- and C-termini will be prepared and use for binding experiments with PIP2 employing spectroscopy methods. The data will be correlated with results from electrophysiology experiments. TRPM channels and their appropriate mutated constructs in putative PIP2 binding sites will be expressed in HEK 293 cells. Homology models of TRPM channels based on known structures of TRPV1, TRPV2 and TRPA1 channels will be studied by means of molecular dynamics simulations for their interactions with PIP2.
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Elucidation of the structure and interactions of glycoconjugates and glycans using vibrational spectroscopy
Grant agency: GACR
Identification number of grant: 16-00270S
Head of the project: Ing. Jakub Kaminský, Ph.D. (Institute of Organic Chemistry and Biochemistry AS CR, v.v.i.)
Vice-head of the project:
RNDr. Vladimír Kopecký Jr., PhD.
Grant annotation
Glycans are carbohydrate parts of glycoproteins, glycolipids, or
proteoglycans often determining their function. Glycans involve in
protein folding, protein clearance, cell adhesion, receptor binding,
receptor activation and many other processes. The structural
characterization of glycans in relation to their function is necessary
for their better utilization. However, the scale of tools to study their
structure is limited. We propose an extensive study of glycans using
methods of vibrational optical activity as a suitable tool for
saccharides. The essence of the proposal is to develop new joint
theoretical and experimental methodology for prediction and
interpretation of vibrational optical activity (especially Raman optical
activity) of glycans, their fragments and conjugates and use this
methodology to study their structures, dynamics and interactions. The
project will enhance the usage of methods vibrational optical activity
in the biological studies and beyond, in materials science and
structural studies of glycans.
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Interaction of antibacterial peptides with the model membranes and possibility of the prediction of their biological activity
Grant agency: GACR
Identification number of grant: P208/10/0376
Head of the project: RNDr. Lucie Bednárová, CSc. (Institute of Organic Chemistry and Biochemistry AS CR, v.v.i.)
Vice-head of the project:
RNDr. Vladimír Kopecký Jr., PhD.
Grant annotation
A set of experimental techniques, like circular dichroism (CD), stopped
flow CD, FTIR, Raman spectroscopy, and fluorescence spectroscopy
including fluorescence correlation spectroscopy will be employed to
study the newly isolated antibacterial peptides (AMP) from bees’ venom
and their synthetic analogs. Mechanism of AMPs action will be
investigated by means of interaction with the selected model membranes
mimicking different types of cells. The proposed model systems in
combination with the selected peptides and the used experimental methods
enable understanding mode of action of AMPs. Correlation of the results
with biological tests will be helpful for the design of the peptide
analogues synthesized for medical purpose.
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Testing of the stereospecific binding between cerebrosterol and amyloid beta peptides
Grant agency: GACR
Identification number of grant: 305/09/0457
Head of the project: Ing. Zdena Krištofiková, PhD. (Psychiatrické centrum Praha)
Vice-head of the project:
RNDr. Vladimír Kopecký Jr., PhD.
Grant annotation
Although causes of Alzheimer disease are not known yet in a detail, it
is suggested that amyloid beta peptides play a key role in the
pathogenesis of disease. Toxicity of peptides is probably closely
connected with their ability to ologomerize or to form complexes with
different endogenous substances. A great attention is now focused on the
research od drugs that via binding to amyloid beta peptides can
eliminated their toxic effects and that could be applied in Alzheimer
disease therapy. E.g., the stereospecific binding to naturally occurring
22R-hydroxy-cholesterol decreases peptide toxicity and enantiomer
22S-hydroxycholesterol is not effective. Naturally occurring and
brain-originating 24S-hydroxycholesterol (cerebrosterol) is a very
efficient inhibitor of amyloid beta peptide secretion in experiments in
vitro. Our previous work indicated the decreased actions of
nonaggregated and aggregated peptides when racemic 24-hydroxycholesterol
was added and data suggested a formation of the complex as probable.
However, interaction mechanisms are not known yet in a detail.
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New approaches for study of protein structure by means of vibrational spectroscopy
Grant agency: GAAS
Identification number of grant: KJB101120805
Head of the project: RNDr. Vladimír Kopecký Jr., PhD.
Vice-head of the project:
RNDr. Kateřina Hofbauerová, PhD.
Grant annotation
Determination of the protein structure represents one of the key tasks
of present molecular biology. There is an urgent need for
tree-dimensional structures of proteins for understanding their role in
organisms and efficient drug design. The main goal of the project is
development of the new methodology for estimation of the secondary
structures content of proteins by means of vibrational spectroscopy
(Raman spectroscopy, Raman optical activity, drop deposition-coating
Raman spectroscopy, Fourier-transform infrared spectroscopy and
Fourier-transform infrared spectroscopy with attenuated total
reflection). The large protein reference spectra set for each method
will be measured, present methods will be implemented and new methods
(based on multivariate statistic) with higher information content, e.g.
determination of the number of structure segments, of the protein fold
etc., will be developed. The methods will be freely available and can be
applied in future protein research.
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Structural and binding properties of human alpha(1)-acid glycoprotein
Grant agency: GACR
Identification number of grant: 202/06/P208
Head of the project: RNDr. Vladimír Kopecký Jr., PhD.
Grant annotation
The project is devoted to the study of human alpha (1)-acid glycoprotein
(AGP), also known as orosomucoid. The protein plays an important role in
a blood plasma under pathophysiological conditions, therefore
pharmaceuticals are targeted on it. Nevertheless, its structure and
function remains unknown. The project will be focused on detail study of
structure of AGP and its genetic variants, on the first by means of
physical experimental methods of optical spectroscopy (Raman
spectroscopy, Raman optical activity, infrared and UV-VIS spectroscopy)
and on the latter by means of theoretical calculations, i.e. by computer
modeling of proteins. The binding of ligands (with biomedical usage) to
AGP will be studied as well. The aim of the project is to build
experimentally verified computer models of genetic variants of AGP,
characterize their binding sites in detail and understand the role of
carbohydrate moiety in its biological and structural function. The
project will significantly contribute to understanding of structural and
binding properties of AGP and it enables effective drug design of
pharmaceuticals targeted on this protein.
