Our research group has several interests, the largest effort being to better understand protein structure. Our goal is to explore and understand the underlying physical and chemical principles that determine a protein's secondary, tertiary and quaternary structure as well as how these factors contribute to the overall stability of the structure once it is formed. In the longer term, we want to apply this knowledge to practical purposes such as increasing protein stability and designing protein structures. In order to accomplish these objectives we use the techniques of molecular biology, biophysics, organic and peptide chemistry.
We use the protein staphylococcal nuclease as a model system. In a typical project, we chose a subset of residues and make a series of mutations. We have literally made and characterized hundreds of mutants. For example, we have studied the effects of adding or removing hydrogen bonding capacity in side chains and the effects of increasing the hydrophobicity of solvent exposed residues. In some instances we may follow this by chemical modification, such as when we crosslinked the protein at introduced cysteine residues. More recently, we have constructed the world’s largest mutant library examining the role of side chain packing in stabilizing protein structure. We then characterize the effects of the mutation on protein stability, association and structure by a variety of biophysical techniques. Circular dichroism, NMR and fluorescence spectroscopy, analytical ultracentrifugation, dynamic light scattering, and calorimetry are among the techniques at our disposal. Our group collaborates closely with Dr. Sakon’s to carry out X-ray crystal structure determination of many mutants. Once we have characterized the behavior of the mutant protein we use statistical and modeling procedures to determine the causes of the observed changes from the behavior of the wild-type protein. Our objective is to eventually be able to accurately predict the energetic and structural consequences of any mutational change.
A new focus of research interest has been the oxidation of methionine. This chemical modification of proteins is an apparently important process in human disease and aging. We are interested in it as a possible mechanism for the reversible regulation of protein activity. Of particular interest to us is the effect of the oxidation of Met388 in thrombomodulin, a key protein in the regulation of blood coagulation.
NIH Postdoctoral Fellow, Johns Hopkins University School of Medicine
Ph.D., Massachusetts Institute of Technology