Areas of Interest
Our group studies protein structure using X-ray crystallography in order to understand protein function. We use the three-dimensional structures of proteins to understand molecular mechanisms of biological processes and to develop hypotheses about function, which we test through experiments in our lab or by collaboration. Two current projects illustrate our approach.
Glutamine amidotransferases are complex enzymes that produce ammonia from the amide group of glutamine, and channel the ammonia through an intra-molecular tunnel to a second active site where it is added to a second substrate. Amidotransferases occur in a variety of pathways for biosynthesis of nitrogenous molecules, including purine and pyrimidine bases, cofactors, aminosugars and amino acids. Our crystal structures of glutamine PRPP amidotransferase (purine pathway), GMP synthetase (guanine pathway), imidazole glycerol phosphate synthase (histidine pathway) and other enzymes show that the chemical steps of glutamine hydrolysis and amination occur independently, but that they are coordinated by the protein through a variety of conformational switches. Our analysis of the structures has shown how each enzyme keeps its glutaminase active site in an inactive state until the second substrate is bound and ready to accept the labile ammonia. Currently challenges are to understand the conformational switches used for cross-talk by the distant active sites and to study new amidotransferases that have been detected in genome sequences.
The replication machinery of alphaviruses and flaviviruses also represents a complex catalytic system. These positive-sense RNA viruses encode enzymes for genome replication, RNA capping, RNA unwinding and polyprotein processing. Specific protein-protein complexes, as yet poorly characterized, coordinate the catalytic activities and specify particular RNA or protein substrates at appropriate times in the viral life cycle. Our structural studies of individual enzymes and enzyme complexes aim to elucidate mechanisms of catalysis and of time-dependent activation and specificity.
Honors & Awards
1998-2000 NIH Merit Award
1998-2008 NIH Merit Award
2001 Herbert Newby McCoy Award, Purdue University
2007 AAAS Fellow
A biologist's guide to synchrotron facilities: the BioSync web resource. A. Kuller, W. Fleri, W. F. Bluhm, J. L. Smith, J. Westbrook, and P. E. Bourne (2002) Trends Biochem. Sci., 4, 213-215.
Addition of synthetic lipid to the cytochrome b6f membrane protein-detergent complex enables crystallization.H. Zhang, G. Kurisu, J. L. Smith, and W. A. Cramer, W. (2003). Proc. Natl Acad. Sci. USA, 100, 5160-5163.
Towards understanding the mechanism of the complex cyclization reaction catalyzed by imidazole glycerolphosphate synthase: Crystal structures of a ternary complex and the free enzyme. B. N. Chaudhuri, S. C. Lange, R. S. Myers, V. J. Davisson and J. L. Smith (2003) Biochemistry 42, 7003-7012.
Substrate-Induced changes in the ammonia channel for imidazole glycerol phosphate synthase. R. S. Myers, J. R. Jensen, I. Deras, J. L. Smith and V. J. Davisson (2003) Biochemistry42, 7013-7022.
The purine repressor of Bacillus subtilis: a novel combination of domains adapted for transcription regulation. S. C. Sinha, J. Krahn, B. S. Shin, D. R. Tomchick, H. Zalkin and J. L. Smith. (2003) J. Bact., 185, 4087-4098.
Functional dissection of the Bacillus subtilispur operator site. A. K. Bera, J. Zhu, H. Zalkin and J. L. Smith. (2003) J. Bact., 185, 4099-4109.
Structure of the cytochrome b6fcomplex of oxygenic photosynthesis: tuning the cavity. G. Kurisu, H. Zhang, J. L. Smith and W. A. Cramer. (2003) Science302, 1009-1014.
Crystal structure of imidazole glycerol phosphate dehydratase: duplication of an unusual fold. S. C. Sinha, B. N. Chaudhuri, J. W. Burgner, G. Yakovleva, V. J. Davisson and J. L. Smith (2004) J. Biol. Chem. 279, 15491-15498.
The structure of the RNA-dependent RNA polymerase from bovine viral diarrhea virus establishes the role of GTP in de novo initiation. K. H. Choi, J. M. Groarke, D. C. Young, R. J. Kuhn, J. L. Smith, D. C. Pevear and M. G. Rossmann. (2004) Proc. Natl Acad. Sci. U S A. 101, 4425-4430.
Design, expression, and purification of a Flaviviridae polymerase using a high-throughput approach to facilitate crystal structure determination. K. H. Choi, J. M. Groarke, D. C. Young, M. G. Rossmann, , D. C. Pevear, R. J. Kuhn and J. L. Smith. (2004) Prot. Sci. 13, 2685-2692.
Self-processing cysteine-dependent N-terminal nucleophile hydrolases. J. L. Smith. (2004) in Handbook of Proteolytic Enzymes 2nd Edn, A. J. Barrett, N. D. Rawlings and J. F. Woessner eds., pp. 2050-2052, Academic Press, San Diego.
Structure of the nucleotide complex of PyrR, the pyr attenuation protein from Bacillus caldolyticus, suggests dual regulation by pyrimidine and purine nucleotides. P. Chander, K. M. Halbig, J. K. Miller, C. J. Fields, H. K. S. Bonner, R. L. Switzer and J. L. Smith. (2005) J. Bacteriol. In press.