Research Interests

Biophysical chemistry: Mechanisms of Protein Folding and Function 

My research interests are the molecular mechanisms of protein folding and enzyme function. To understand these processes, it is necessary to characterize the structural transitions in proteins along the folding, binding and catalytic pathways as well as the roles of the individual amino acid residues. For this purpose, we are using isotopically-edited infrared (IR) spectroscopy to selectively probe the structure of particular regions of the studied proteins. Time-resolved IR, in combination with pulsed laser temperature jump, which can trigger the folding or binding process within nanoseconds, will allow tracking and characterization of short-lived kinetic intermediates.

Protein folding

Our protein folding studies initially focus on the simplest proteins: the helix-turn-helix motifs. Characterizing the folding mechanism of simple structural motifs is necessary for understanding folding of more complex protein structures and will be an important starting point for future investigations of folding of larger proteins.

 

 

The ¹³C isotopically labeled proteins are synthesized in my laboratory using FMOC solid phase peptide synthesis. Chemical synthesis also allows incorporation of specific amino acid mutations into the protein sequence. The mutations (“protein engineering”) are used to probe the roles of the individual amino acids in formation and stabilization of the native protein structure, but also of the intermediate and transition structures along the folding pathways.

To help interpret the infrared spectroscopic data as well as design optimal isotopical labeling schemes, we carry out quantum mechanical calculations of IR spectra for model peptides using density functional theory. IR spectra of much larger structures, such as small proteins are then calculated by transferring the vibrational parameters obtained from the first principles. 

Enzyme-ligand binding

Specific protein dynamics is believed to be crucial for binding substrates and enzymatic catalysis. We will apply the same battery of techniques used for studies of the protein folding, namely the isotopically-edited IR and protein engineering approaches, to investigate the structural dynamics of simple enzymes during ligand binding. Our goals are to characterize the enzyme conformational transitions in terms of local protein structural dynamics during binding and investigate the effects of perturbations to the chain flexibility and residue interactions in the vicinity of the enzyme active site on binding by mutations.

 
Enzymatic catalysis in non-aqeuous environments

Although aqueous solution is the natural environment of proteins, enzymes can not only retain most of their catalytic function in non-aqueous media, but the solvent environment can regulate many important properties of enzymes, such as substrate specificity. One of our objectives is to study the catalytic efficiency of enzymes in different solvent environments. Correlation of these data with the protein stability and flexibility under different solvent conditions will lead to a better mechanistic understanding of enzymatic activity and of the unusual enzyme properties in organic solvents. Such insights are crucial for further development of enzymatic systems suitable for applications as catalysts for processes that are not compatible with aqueous solutions.