Background:

• Research Associate, The Scripps Research Institute, 1991-95
• NSF Early Career Development Award, 1996-2001
• Indiana University Outstanding Junior Faculty Award, 1999-2000

We utilize multidimensional NMR spectroscopy in combination with molecular biology/biochemistry approaches to understand the influence of structure and dynamics on protein function.

Structure-activity relationships of chemokines. Chemokines are small soluble proteins that mediate inflammatory responses by stimulating the migration of leukocytes. Inhibition of chemokines is an attractive and potentially general approach to treatment or prevention of inflammatory diseases. Our work focuses on the eotaxin subgroup chemokines which trigger the accumulation of eosinophils in allergic diseases (e.g., asthma) and in parasitic infections. We are using NMR to determine the three-dimensional structures of the chemokines, site-directed mutagenesis along with functional assays to identify the regions of the chemokines important for receptor activation, and peptide models to probe the nature of the chemokine-receptor interactions. These studies will further our understanding of the biochemistry of inflammation and may lead to the development of chemokine inhibitors as anti-inflammatory drugs.

Protein dynamics. Folded protein chains retain a significant amount of mobility (conformational entropy). Changes in this conformational entropy may occur upon variation of environmental conditions or upon ligand binding, giving rise to changes in the stability of the protein or the protein-ligand complex. We are using 2D NMR methods to measure the flexibility of backbone NH groups in proteins. We have found that association of a mouse pheromone with its binding protein leads to an increase in backbone mobility of the protein, thus stabilizing the complex. We have also found that variations of protein dynamics among protein mutants may correlate with their differences in thermal stability. In addition, temperature dependence studies of backbone motions suggest a role for backbone heat capacity in the stabilization of thermophilic proteins. Recently, we have compiled and analyzed a database of NMR-derived dynamics data to identify relationships between protein structure and dynamics. Together, these studies are allowing us to develop an understanding of the factors that influence protein entropy. Incorporation of these principles into molecular modeling will improve the predictive value of these methods for drug design.

Selected Publications:

"Active monomeric and dimeric forms of Pseudomonas putida glyoxalase I: evidence for 3D domain swapping," with A. P. Saint-Jean, K. R. Phillips, and D. J. Creighton. Biochemistry, 37, 10345 (1998).

"NMR mapping of the recombinant mouse major urinary protein I binding site occupied by the pheromone 2-sec butyl-4,5-dihydrothiazole," with L. Zidek, S. M. Lato, A. D. Ellington, and M. V. Novotny. Biochemistry, 38, 9850 (1999).

"Backbone dynamics of the human CC-chemokine eotaxin," with J. Ye and K. L. Mayer. J. Biomolecular NMR, 15, 115 (1999).

"Increased protein backbone conformational entropy upon hydrophobic ligand binding," with L. Zidek and M. V. Novotny. Nature Structural Biology, 6, 1118 (1999).

"Three-dimensional structure of chemokines," with K. L. Mayer. In Chemokines in Allergic Disease, pp. 67-94, M. Rothenberg, ed. New York: Marcel Dekker, 1999.

"Relationships between protein structure and dynamics from a database of NMR-derived backbone order parameters," with J. L. Goodman and M. D. Pagel. J. Mol. Biol., 295, 963 (2000).

"NMR solution structure and backbone dynamics of the CC chemokine eotaxin-3," with J. Ye, K. L. Mayer, and M. R. Mayer. Biochemistry, 40, 7820 (2001).

"Identification of receptor binding and activation determinants in the N-terminal and N-loop regions of the CC chemokine eotaxin," with M. R. Mayer. J. Biol. Chem., 276, 13911 (2001).

"NMR relaxation studies of the role of conformational entropy in protein stability and ligand binding." Acc. Chem. Res., 34, 379 (2001).

"Comparison of protein backbone entropy and b-sheet stability: NMR-derived dynamics of protein G B1 domain mutants," with S. Gupta, N. Snyder, and L. Regan. J. Am. Chem. Soc., 123, 185 (2001).