Protein Structure and Function



	Philip J. Fay, Ph.D. The severe, inherited bleeding disorder, Hemophilia A, results from defects or deficiency in the plasma protein, factor VIII. This protein serves as a cofactor for the serine protease, factor IXa, in the set of reactions referred to as the blood coagulation cascade that ultimately yields a clot to prevent blood flow. Our laboratory investigates the physical and biochemical properties of the labile factor VIII protein as well as its interactions with factor IXa and other components of the cascade. Our goals are to determining molecular mechanisms for the cofactor activity of this protein.

	Barry M. Goldstein, M.D., Ph.D. The structure of a drug, ligand, target enzyme or receptor at atomic resolution ultimately leads to an understanding of its functional properties. Our laboratory uses macromolecular crystallography aimed at determining protein-ligand interactions of molecules with either medical or biotechnology applications. Our studies build on understanding basic enzyme mechanisms and have applications in the rational design of anti-tumor and antiviral drugs and in the design of enzymes to synthesize chemicals in an environmentally friendly way.

	Fred K. Hagen, Ph.D. The formation of adult tissues and organ systems requires the integrated interactions of multiple cell types and cell surface molecules. Glycosyltransferases modify cell surface and secreted molecules and in a few cases are known to influence developmental processes like axon guidance, cell migration, cell signaling, and cell adhesion. A comprehensive functional genomics study on the role of glycosyltransferases in development is being complemented with detailed biochemical characterization of specific glycosyltransferases that are essential for development.

	Jeffrey J. Hayes, Ph.D. During the cell cycle, genomic DNA assembles into a compact, higher ordered chromatin fiber. The organization of chromatin structure is regulated in a gene-specific fashion and integrated with the machinery that controls transcription. Our laboratory studies defined protein-DNA interactions, protein modification and mutations that influence chromatin structure and gene regulation. Specific mutations of histone proteins are being used to probe protein domains involved in site-specific contacts between DNA and nucleosomes.

	Clara Kielkopf, Ph.D. Noncoding sequences interrupt almost all human genes, and must be removed from pre-mRNAs by splicing before translation into proteins. Our laboratory seeks to understand the structural, thermodynamic, and kinetic characteristics driving protein-RNA and protein-protein interactions during identification and pairing of the the appropriate pre-mRNA splice sites. Given that errors in pre-mRNA splice site identification account for more than 50% of human genetic diseases, and are frequently associated with cancers and leukemias, biophysical maps of the key molecular interactions provide foundations for new therapeutic approaches.

	Mahin D. Maines, Ph.D. Neurons are susceptible to oxygen free-radicals, and oxidative stress can induce neuronal cell death. Protection from neuronal oxidative damage is mediated by the heat shock family 32 protein, heme oxygenase-1. We are using a transgenic mouse model system to study the gene regulation and protective properties of heme oxygenase isozymes. Furthermore, our studies are also extending to the investigation of the reaction products of the heme oxygenases, which produce signaling molecules and regulate the expression of a cascade of genes.

	Terry Platt, Ph.D. Termination of transcription and its degree of coupling to mRNA 3'-end processing is an important aspect of gene expression in both prokaryotic and eukaryotic organisms. We are studying E. coli and yeast as paradigms for these classes, to elucidate the molecular mechanisms governing the formation of mature messenger RNA 3' ends by the transcriptional and processing machinery. Our approaches combine genetic and biochemical techniques to examine the nature of the termination sites in the DNA or RNA, and the proteins and activities that regulate their function.

	Alan E. Senior, Ph.D. Cancer chemotherapy often fails because tumors develop resistance to multiple drugs simultaneously. The culprit is multidrug-resistance protein (MDR), a transport protein that uses ATP to pump drugs out of the cell. The long-term goal or our research is to learn ways to disable MDR protein in cancer cells. We are also using molecular genetics in bacteria in a biophysical study of the mechanism of action of ATP-linked membrane transporters.

	Harold C. Smith, Ph.D. mRNA is not only spliced, but also edited to produce a final processed message. mRNA editing is regulated developmentally in a tissue-specific manner and can be significantly altered by diet, hormonal changes, and toxins such as ethanol. We are studying the regulation of editing at the level of cell signaling and trafficking of the editing factors between the cytoplasm and the nucleus (ultimately the intracellular site where mRNA editing takes place). Animal model systems and whole cell microinjections analyses are being used to determine the molecules and mechanisms involved in apoB mRNA editing.

	Joseph E. Wedekind, Ph.D. The C6666 to U modification of apoB mRNA represents the archetypal RNA editing reaction. This phenomenon requires a Zn-dependent cytidine deaminase, APOBEC-1, which resides at the heart of a multi-protein particle dubbed the nuclear 27S editosome. Other essential proteins include an RNA binding factor (ACF), and a 240 kDa assembly factor. We have undertaken X-ray crystallographic structure determinations of whole 27S particles, as well as isolated proteins in order to elucidate the editosome mechanism. Ultimately this work will establish a working model for mammalian RNA editing.

	Yi-Tao Yu, Ph.D. Pre-mRNA splicing is essential for the appropriate excision of introns and expression of mature gene products. The splicing reaction occurs in the spliceosome, a massive complex containing five small nuclear RNAs (snRNAs) and a large number of proteins. Interestingly, the five snRNAs are all post-transcriptionally modified. Our research focuses on spliceosomal snRNA modifications and their roles in pre-mRNA splicing. Furthermore, we use a combined approach of molecular biology and cell biology to determine how and where within the cell the spliceosomal snRNAs are modified.