Founded in 1975 as an independent research institution, now part of Vilnius University, the Institute of Biotechnology strives to maintain the high standards of excellence in scientific endeavour, research training and technological advance with its main focus in a broadly defined field of molecular biotechnology including nucleic acid and protein technologies, bioinformatics, molecular diagnostics, drug design, next generation epigenomic and gene editing technologies. The Institute provides an interface between advanced education, basic research and technological development for the economic and social benefit of Lithuania.


Department of Biothermodynamics and Drug Design • Research projects




Several protein targets have been selected for the investigation of protein – ligand binding thermodynamics and the design of novel ligands with desired properties. A family of human carbonic anhydrases, heat shock proteins, and several signal-tranducing proteins were chosen as anticancer drug targets.

Novel methods and thermodynamic approaches are being used and developed in the laboratory. Detailed thermodynamic description of natural compound – protein interaction provides clues to improved ligand affinity and specificity. In addition to the Gibbs free energy, enthalpy, entropy, and the heat capacity, the laboratory studies the volume and compressibility of the protein – ligand interactions.

The laboratory has been recently invited to write a review chapter on the thermal shift assay in Royal Society of Chemistry “Protein – Ligand Binding” series book (Cimmperman and Matulis, 2010).

Carbonic anhydrases as anticancer drug targets

Carbonic anhydrases (CAs), a group of zinc containing enzymes, are involved in numerous physiological and pathological processes, including gluconeogenesis, lipogenesis, ureagenesis, tumorigenicity and the growth and virulence of various pathogens. In addition to the established role of CA inhibitors as diuretics and antiglaucoma drugs, it has recently emerged that CA inhibitors could have potential as novel anti-obesity, anticancer, and anti-infective drugs (Supuran, 2007). CAs catalyse a simple reaction – the conversion of CO2 to the bicarbonate ion and protons. There are 12 catalytically active CA isoenzymes in humans. A number of CA inhibitors, mostly unsubstituted sulfonamides, have already been designed. However, most present inhibitors are insufficiently selective for target CA isozymes, such as hCAIX and hCAXII, anticancer targets.

Here at the LBDD we have cloned and purified most soluble CAs and truncated versions of CAs with removed transmembrane domains. Laboratory participated in the characterization of hCA IX (Hilvo et al, 2008). The organic synthesis team together with collaborators designed and synthesized over 200 novel compounds that bind CAs with submicromolar affinity. Several novel groups of CA inhibitors exibited high affinity and appreciable selectivity towards selected CA isozymes (Dudutiene et al. 2007; Baranauskiene et al. 2010; Sudzius et al. 2010; Capkauskaite et al. 2010).

Figure. Left panel. View of compound 3m bound in the active center of hCA II. The Zn atom, His94, His96, and His119 are shown as transparent, inhibitors are shown in orange. Right panel. Superposition of the hCA II–indapamide (magenta), hCA II-chlorthalidone (cyan) with hCA II – 3m (orange). Zinc ion is shown as pink sphere, His94, His96, His119, and protein secondary structures are in green. The crystal structure was solved in collabora-tion with dr. Saulius Gražulis group, Laboratory of Protein-DNA Interactions (Capkauskaite et al, 2010).

Inhibition of Hsp90 chaperone

Heat shock protein 90 (Hsp90) is a molecular chaperone that is responsible for the correct folding of a large number of proteins. Client proteins of Hsp90 include many overexpressed oncogenes that are critical for the transformed phenotype observed in tumours.

Our laboratory is interested in the mechanism of Hsp90 action and the thermodynamics of inhibitor binding. Thermodynamics of a natural compound radicicol binding to human Hsp90 alpha and beta isozymes and yeast Hsc82 was studied by isothermal titration calorimetry, thermal shift assay, and the pressure shift assay. These studies provided an unusual and detailed picture of Hsp90 inhibitor binding energetics. Radicicol bound Hsp90 with exceptionally large exothermic enthalpy and volume of binding (Zubriene et al 2009, Zubriene et al. 2010, Toleikis et al, submitted).

A novel group of inhibitors has been designed and synthesized that is similar to radicicol, a well known natural inhibitor of Hsp90.


Studies of Protein Denaturation by High Pressure

Pressure studies yield a variety of thermodynamic parameters that are unavailable by other methods, such as molar volume, expansion coefficient, and compressibility of the proteins in the native and denatured states. Ligand binding affects protein temperature and pressure stability. The pressure stability information yields novel methods of determining ligand binding equilibria.



The LBDD is seeking to license out the compounds described in patent applications. The LBDD is also interested in the collaborations where our expertise in the determination of compound – protein binding thermodynamics and recombinant protein stability characterization could be applied. Protein – ligand binding constants and protein thermal stability profiles at hundreds of conditions may be determined in a single experiment by consuming microgram quantities of protein.