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Our lab is using biophysical methods and molecular biology to study protein-nucleic acid interactions and macromolecular assembly. We are particularly interested in mechanisms of assembly and how kinetics and equilibria influence the final composition of assembly complexes and ultimately biological function. Experiments are aimed at determining the detailed function of proteins and how they recognize nucleic acids. These processes are studied by steady-state and life-time fluorescence measurements, circular dichroism, rapid scanning stopped-flow, and site-directed mutagenesis. Detailed quantitative measurements of these biological processes lead to elucidation of molecular mechanisms and how these processes can be regulated.
Plant Viral Protein Synthesis
Plant viral diseases affect a significant number of food crops world-wide and can have severe impact both on economic conditions and food supply. Protein synthesis is a key step in viral infection yet is not well understood. This research will increase our understanding of plant viral protein synthesis and how this important biochemical process can be regulated. This information can potentially be used to shut off viral protein synthesis either through genetic engineering or biochemical agents. Further, viral protein synthesis in general is simpler than host cell protein synthesis so there is potential to use the information from this research to develop systems to produce desired proteins which are nutritionally beneficial, antibodies, or have other economic uses. -more-
Assembly of Transcription Initiation Complex
Myc/Max/Mad is an important biological system both as a model for protein-DNA interactions and assembly and because of the cancer potential associated with these proteins. Deregulation of Myc, which has strong oncogenic potential, is associated with numerous types of human cancer including lymphoid malignancies, lung cancer, breast cancer and colon cancer. This project will investigate the interactions of three members of the b/HLH/Z family of transcription factors, Myc, Max, and Mad. In order to be functional, Myc interacts with Max and acts as a transcriptional activator with strong oncogenic potential. Max also dimerizes with Mad, however this heterodimer acts as a repressor of transcription and a tumor suppressor. Transcription requires a very complex assembly of a large number of proteins on the appropriate DNA site. This macromolecular assembly is modulated by many interactions such as protein-protein interactions, phosphorylation, cooperative or anticooperative binding, etc. This research is expected to increase our understanding of gene function, provide a basis for elucidating the effects of other transcription factors, and identify new targets for control of a multitude of human cancers. -more-
Cap binding proteins and decapping enzymes, viral protein-RNA interactions, plant anti-viral proteins, DNA interacting proteins
National Science Foundation, NIH, Promega Biotech, and Life Technologies
Browning, K. S., Goss, D.J. and Gallie, D. (l998) "The Translational Machinery of Plants" A Look Beyond Transcription. American Soc. of Plant Physiologists. 68-84.
Ruud, K., Kuhlow, C. Goss, D.J. and Browning K.S. (l998) "Identification and Characterization of a Novel Cap Binding Protein from Arabidopsis thaliana. J. Biol. Chem. 273, 10325-10330.
Wei, C-C., Balasta, M.L., Ren, J. and Goss, D. J. (l998) "Wheat germ Poly(A) Binding Protein Enhances the Binding Affinity of Eukaryotic Initiation Factor 4F and (iso)4F for Cap Analogues. Biochemistry 37, 1910-1916.
Bi, X., Ren, J., and Goss, D.J. (2000) "Wheat Germ Translation Initiation Factor eIF4B Affects eIF4A and eIFiso4F Helicase Activity by Increasing the ATP binding Affnity of eIF4A" Biochemistry 39, 5758-5765.
Bi, X. and Goss, D.J. (2000) "Wheat Germ Poly A binding Protein Increases the ATPase and the RNA Helicase Activity of Translaion Initiation Factors eIF4A, eIF4B, and eIF-iso4F" J. Biol. Chem. 275, 17740-17746.
Bi, X. and Goss, D.J. (2000) "Kinetic Proofreading Scanning Model for Eukaryotic Translation Initiation: the Cap and Poly A tail Dependency of Translation" J. Theor. Biology 207, 145-157.
Luo, Y. and Goss, D.J. (2001) "Homeostasis in mRNA Initiation. Wheat Germ Poly(A)-binding Protein Lowers the Activation Energy Barrier to Initation Complex Formation" J. Biol. Chem. 16, 43083-6.
Scheper, G.C. van Kollenburg, B. Hu, J. Luo, Y., Goss, D.J. and Proud, C.G. (2002) Phosphorylation of eukaryotic initiation factor 4E markedly reduces its affinity for capped mRNA. J. Biol. Chem. 277, 3303-3309.
Spivak-Kroizman, T., Friedland, D.E., De Staercke, C., Gernert, K. M., Goss, D.J. and Hagedorn, C.H. (2002) Mutations in the S4-H2 loop of eIF4E which increase the affinity for m7GTP. FEBS Letters 25892, 1-6.
Khan, M.A. and Goss, D.J. (2004) "Phosphorylation States of Translational Initiation Factors Affect mRNA Cap Binding in Wheat" Biochemistry 43, 9092-9097.
Khan, M.A. and Goss, D.J. (2005) "Translation Initiation Factor (eIF4B) Affects the Rates of binding of the mRNA m7G cap Analogue to Wheat Germ eIFiso4F and eIFiso4F-PABP" Biochemistry 44, 4510-4516.
Friedland, D.E., Wooten, W.N.B., LaVoy, J.E., Hagedorn, C.H. and Goss, D.J. (2005) "A Mutant of Eukaryotic Protein Synthesis Initiation Factor eIF4EK119A has an Increase Binding Affinity for both m7G Cap Analogues and eIF4G Peptides" Biochemistry 44, 4546-4550.