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SCOTT STROBEL

Riboswitches. Riboswitches are noncoding RNAs that regulate the expression of genes in response to the presence of small molecules. We use biophysical and biochemical techniques aided by synthetic organic chemistry to study the structures and functions of these important RNAs. We have biochemically characterized and solved the X-ray structures of several riboswitch classes: the glycine riboswitch, which binds its ligand cooperatively; two classes of c-di-GMP riboswitches, which bind an important second messenger and function in cell signaling; and the glmS ribozyme, which controls gene expression by self-cleaving in the presence of its ligand. We are currently pursuing several approaches in order to more fully understand these macromolecules. Efforts include determining the atomic resolution structures of several new riboswitch classes as well as alternative complexes, investigating intramolecular interactions in the glycine riboswitch to understand the molecular basis of cooperativity in an RNA system, designing and testing ligand analogs to probe small molecule-RNA interactions for the design of potential drug candidates and performing in vivo studies of riboswitch variants to determine their regulatory mechanisms.

Protein Synthesis. Protein synthesis in all organisms is catalyzed by the ribosome, a large complex of RNA and protein. Crystallographic studies have revealed that the peptidyl transferase center, where the chemical reactions of protein synthesis take place, is composed exclusively of RNA. Therefore, this ancient and essential process is catalyzed not by protein but by RNA: the ribosome is a ribozyme. We are applying biophysical techniques with the help of synthetic chemistry to determine how the ribosome catalyzes this fundamental reaction of biology. We have determined the transition state for peptide bond formation using a combination of kinetic isotope effects, linear-free energy relationships, and transition state analogs. We are extending our studies to peptide release, the final reaction of protein synthesis. Again aided by synthetic chemistry, we are using kinetic isotope effects and transition state analogs to determine the structure of the transition state for this reaction and the relative positioning of ribosomal functional groups. We are also studying a third chemical reaction on the ribosome - the cleavage of mRNA by the ribosome-dependent endonuclease RelE. RelE is structurally similar to other endoribonucleases, but it lacks conserved catalytic residues and it is only active in the context of the ribosomal A-site. We are using biochemical and genetic approaches to establish how RelE residues and the ribosome each contribute to this cleavage reaction.

Hydrocarbon production by endophytic fungi. Another research direction of the lab is the study of secondary metabolism of endophytes with particular focus on the production of and resistance to molecules with applications as biofuels. Endophytes are attractive targets for biofuel production because they produce many diverse products and most naturally digest cellulose, offering a potential production pathway that is carbon neutral. Our strategy for identifying these products and their pathways includes (i) natural product purification and characterization, (ii) elucidation of the biosynthetic pathways of these products using metabolic labeling (iii) genomic, transcriptomic and metabolomic correlation of expression with production of metabolites of interest, and (iv) validation of genetic pathways in heterologous systems. In addition to known fungal systems, the lab has access to a large number of fungal endophytes that are taxonomically novel at the level of genus or higher through the course “Rainforest Expedition and Laboratory.”

K. D. Smith, C. A. Shanahan, E. L. Moore, A. C. Simon, S. A. Strobel, Structural basis of differential ligand recognition by two classes of c-di-GMP binding riboswitches, Proc. Natl. Acad. Sci. U.S.A. 108, 7757-7762 (2011).

E. B. Butler, Y. Xiong, J. Wang and S. A. Strobel. Structural basis of cooperative ligand binding by the glycine riboswitch. Chem Biol. 18, 293-298 (2011).

M. A. Griffin, D. J. Spakowicz and S. A. Strobel. Volatile organic compound production by organisms in the Ascocoryne genus and a reevaluation of myco-diesel production by NRRL 50072. Microbiology 156, 3814-3829 (2010).

K. D. Smith, S. V. Lipchock, T. D. Ames, R. R. Breaker and S. A. Strobel, Structural basis of ligand binding by a c-di-GMP riboswitch Nature Struc. Molec. Biol.16, 1218-1223 (2009).

D. A. Hiller, V. Singh, M. Zhong and S. A. Strobel, A two-step chemical mechanism for ribosome-catalyzed peptide bond formation, Nature 476, 236–239 (2011).

Last Updated 8/16/11 myc