My laboratory is primarily interested in biomolecular NMR spectroscopy, protein structural biology, and drug design. Recent work has centered around structure/function investigations and the characterization of motional dynamics in proteins and the development of computational methods and NMR methodologies for macromolecular structure refinement. Some of these investigations involve cross-disciplinary collaborative interactions with other research groups at UAB and elsewhere.
Protein Structural Biology: Current work is aimed at understanding the structural biology of the capsid proteins of the HIV-1 virus and other retroviruses. The wild type capsid protein of HIV-1 is particularly challenging to study by 3D/4D-NMR because of its monomer-dimer equilibrium in solution and its tendency to form oligomers. It was also difficult to study by x-ray crycoverstallography because of its flexibility. Thus, we have utilized a double mutant of the HIV-1 CA (these mutations abolish its dimerization and infectivity, but preserve most of the properties of the wild type protein), and determined its detailed solution structure by 3D-NMR spectroscopy. The CTD domain in this full-length protein shows some differences from the previously published crystal and NMR structures of the wild type dimer structures. These differences give an insight about the mechanism of dimerization of the wild type protein. The structure of the monomeric mutant HIV-1
CA will aid in the development of novel inhibitors that interfere with the HIV-1 life cycle at the level of assembly of the mature and immature capsids. It will also aid the development of inhibitors that interfere with the interaction of capsid protein with host cell proteins exploited by the virus in its replication cycle. We are also undertaking NMR structural studies to determine the role of calmodulin in Fas-mediated apoptosis of HIV-1 infected cells. A knowledge of the detailed three dimensional structures of calmodulin with cellular targets might lead to the development of calmodulin-antagonists that may have a therapeutic potential.
Methodological Development: Our laboratory has an active interest in developing NMR-based structure-refinement procedures. In collaboration with Dr. Istvan Sugar at the Mt. Sinai Medical Center, New York, our laboratory has developed a variable target function based intensity-restrained global optimization procedure (VARTIGO) for refining the three-dimensional structures of proteins using NOESY data. We have developed an alternative method involving Metropolis Simulated Annealing (MSA) refinement of dihedral angles against experimental
NOESY intensities. Another major project in our laboratory involved the development of a Complete Relaxation and Conformational Exchange Matrix (CORCEMA) procedure for interpreting the NOESY spectra of interacting molecules such as complexes of reversibly forming ligand-receptor complexes. The CORCEMA theory is very general and is applicable over a wide range of udp_gal_gtbdissociation constants for the complex (weak binding to tight binding). It incorporates explicitly all the pertinent protons of the interacting pair, and can account for the effect of motional dynamics in the complexes (e.g., hinge-bending motions in enzymes) on the NOESY. More recently, we have extended the CORCEMA theory to the saturation transfer difference (STD)-NMR experiment. This theory (CORCEMA-ST) is particularly useful in determining the bound conformations of reversibly binding low molecular weight lead compounds recognized by a target protein. Such information in turn can lead to structure-based design of new drugs. The CORCEMA-ST program is currently being utilized by over 82 different research groups including three major pharmaceutical companies in five different continents in their drug-design and developmental work. The CORCEMA and CORCEMA-ST programs may be obtained by contacting Dr. Krishna.
This theory (CORCEMA-ST) is particularly useful in determining the bound conformations of reversibly binding low molecular weight lead compounds recognized by a target protein. Such information in turn can lead to structure-based design of new drugs. The CORCEMA-ST program is currently being utilized by over 75 research groups including three major pharmaceutical companies in five different continents in their drug-design and developmental work. The CORCEMA and CORCEMA-ST programs may be obtained by contacting Dr. Krishna.
Central Alabama High-Field NMR Facility: With support from an NCRR High End Instrumentation Grant, NCI ARRA Supplement to the UAB Cancer Center, the UAB Administration (SOM, OVPRED, and CAS), the Cancer Center, and various Departments and Centers, we have established the Central Alabama High-Field NMR Facility equipped with a state-of-the-art 850 MHz NMR system (with a cryoprobe) as the center piece, along with several lower field NMR systems. This Facility will support the research programs of faculty members at UAB as well as at the neighboring institutions. This facility is located on the 1st floor of the UAB Department of Chemistry (14th street).
