University of Illinois
3107 IGB, MC-195, Box B-4
1206 W. Gregory
Urbana, IL 61801
Professor John A. Gerlt attended Michigan State University where he received his B.S. in Biochemistry in 1969. He received his Ph.D. in Biochemistry and Molecular Biology from Harvard University in 1974. After postdoctoral studies at the National Institutes of Health in 1974-75, he held faculty positions at both Yale University and the University of Maryland before joining the Illinois faculty in 1994. In 2003, he was the recipient of the Repligen Corporation Award in Chemistry for Biological Processes from the Division of Biological Chemistry of the American Chemical Society His research interests are in mechanistic enzymology.
predicting and discovering new enzymatic functions, metabolites, and metabolic pathways; mechanistic enzymology; unnatural mutagenesis
The availability of complete genome sequences permits analyses of the strategies by which Nature can redesign existing enzymes to catalyze diverse reactions. Using tools such as sequence analyses, recombinant DNA methods, enzyme kinetics, and physical organic chemistry, we study three groups of enzymes whose members catalyze different reactions that diverged from a common ancestor.
The members of the enolase superfamily share the ubiquitous β/α-barrel fold. High-resolution structures reveal a conserved binding site for an essential divalent metal ion that stabilizes an enolate anion intermediate. The essential functional groups are located at the ends of the eight β-strands in the pseudosymmetric barrel. Using the paradigm that the reactions catalyzed by members of this superfamily are initiated by abstraction of the α-proton of a carboxylate anion, we have predicted the functions of unknown proteins discovered in genome projects. We are using directed evolution to explore the functional restrictions available to members of this superfamily, aiming to obtain novel catalysts for new reactions.
The members of the crotonase superfamily share an α+ β fold that provides a conserved oxyanion hole used to stabilize enolate anion intermediates derived from coenzyme A esters. In contrast to the enolase superfamily, the positions of the essential functional groups in this superfamily cannot be restricted to known positions within the structure. Therefore, we are studying diverse members to delineate the structural basis for catalytic diversity. For example, we are elucidating the mechanisms of the reactions catalyzed by dihydroxynaphthoyl CoA synthase and 2-ketocyclohexanecarboxyl CoA hydrolase that catalyze the formation and cleavage of carbon-carbon bonds, respectively. By studying this pair of homologous enzymes together, we expect to better define the mechanisms of both reactions.
The members of the orotidine 5'-phosphate decarboxylase "suprafamily" also share the β/α-barrel fold. However, in contrast to the enolase superfamily, these enzymes use conserved functional groups to catalyze reactions involving distinct mechanisms. We are studying enzymes that catalyze aldol and β-ketoacid decarboxylation reactions to better understand this mechanistic plasticity. We will use directed evolution to explore the consequences of this functional plasticity on the design of new enzymes.
Additional Campus Affiliations
Professor Emeritus, Biochemistry
Divergent Evolution in the Enolase Superfamily: Strategies for Assigning Functions, J. A. GERLT, P. C. Babbitt, M. P. Jacobson, and S. C. Almo, J. Biol. Chem. 2012, 287, 29-34.
Homology Models Guide Discovery of Diverse Enzyme Specificities Among Dipeptide Epimerases in the Enolase Superfamily, T. Lukk. A. Sakai, C. Kalyanaraman, S. D. Brown, H. J. Imker, L. Song, A. A. Fedorov, E. V. Fedorov, R. Toro, B. Hillerich, R. Seidel, Y. Patsklvsky, M. V. Vetting, S. K. Nair, P. C. Babbitt, S. C. Almo, J. A. GERLT, and M. P. Jacobson, Proc. Nat. Acad. Sci USA 2012, 109, 4122-4127.
OMP Decarboxylase: Phosphodianion Binding Energy is Used to Stabilize a Vinyl Carbanion Interrmediate, B. Goryanova, T. L. Amyes, J. A. Gerlt, and J. P. Richard, J. Amer. Chem. Soc. 2011, 133, 6545-6548.
Mechanism of the Orotidine 5’-Monophosphate Decarboxylase Reaction: Importance of Residues in the Orotate Binding Site, V. Iiams, B. J. Desai, A. A. Fedorov, E. V. Fedorov, S. C. Almo, and J. A. GERLT, Biochemistry 2011, 50, 8497-8507.
The Enzyme Function Initiative, J. A. GERLT, K. N. Allen, S. C. Almo, R. N. Armstrong, P. C. Babbitt, J. E. Cronan, D. Dunaway Mariano, H. J. Imker, M. P. Jacobson, W. Minor, C. D. Poulter, F. M. Raushel, A. Sali, B. K. Shoichet, and J. V. Sweedler, Biochemistry 2011, 50, 9950-9962.
Activation of R235A Mutant Orotidine 5'-Monophosphate Decarboxylase by the Guanidinium Cation: Effective Molarity of the Cationic Side Chain of Arg-235, S. A. Barnett, T. L. Amyes, B. M. Wood, J. A. Gerlt, and J P. Richard, Biochemistry 2010, 49, 824-826.
Conformational Changes in Orotidine 5’-Monophosphate Decarboxylase: “Remote” Residues that Stabilize the Active Conformation, B. M. Wood, T. L. Amyes, A. A. Fedorov, E. V. Fedorov, A. Shabila, S. C. Almo, J. P. Richard, and J. A. Gerlt, Biochemistry 2010, 49, 3514-3516.
Product Deuterium Isotope Effects for Orotidine 5’ Monophosphate Decarboxylase: Effect of Changing Substrate and Enzyme Structure on the Partitioning of the Vinyl Carbanion Reaction Intermediate, K. Toth, T. L. Amyes, B. M. Wood, K. Chan, J. A. Gerlt, and J. P. Richard, J. Amer. Chem. Soc. 2010, 132, 7018-7024.
Enzyme (Re)Design: Lessons from Natural Evolution and Computation, J. A. GERLT and P. C. Babbitt, Curr. Opin. Chem. Biol. 2009, 13, 10-18.
Mechanism of the Orotidine 5’ Monophosphate Decarboxylase Catalyzed Reaction: Effect of Solvent Viscosity on Kinetic Constants, B. M. Wood, K. K. Chan, T. L. Amyes, J. P. Richard and J A. GERLT, Biochemistry, 2009, 48, 5510-5517.
Mechanism of the Orotidine 5’ Monophosphate Decarboxylase Catalyzed Reaction: Evidence for Substrate Destabilization, K. K. Chan, B. McKay Wood, A. A. Fedorov, E. V. Fedorov, T. L. Amyes, J. P. Richard, S. C. Almo, and J. A. GERLT, Biochemistry, 2009, 48, 5518-5531.
Acetoacetate Decarboxylase: Hydrophobics, Not Electrostatics (News & Views), J. A. GERLT, Nature Chem. Biol. 2009, 5, 454-455.
The Relationship between Active Site Loop Size and Thermodynamic Activation Parameters for Orotidine 5’-Monophosphate Decarboxylase from Mesophilic and Thermophilic Organisms, K. Toth, T. L. Amyes, B. M. Wood, K. K. Chan, J. A. GERLT, and J. P. Richard, Biochemistry 2009, 48, 8006-8013.
Computation-Facilitated Assignment of Function in the Enolase Superfamily: A Regiochemically Distinct Galactarate Dehydratase from Oceanobacililus iheyensis, J. R. Rakus, C. Kalyanaraman, A. A. Fedorov, E. V. Fedorov, F. P. Mills Groninger, R. Toro, J. Bonanno, K. Bain, J. M. Sauder, S. K. Burley, S. C. Almo, M. P. Jacobson, and J. A. GERLT, Biochemistry 2009, 48, 11546-11558.
Gerlt, J. A. (2021). Evolution of Enzyme Function and the Development of Catalytic Efficiency: Triosephosphate Isomerase, Jeremy R. Knowles, and W. John Albery. Biochemistry, 60(46), 3529-3538. https://doi.org/10.1021/acs.biochem.1c00211
Li, Q., Zallot, R., Mactavish, B. S., Montoya, A., Payan, D. J., Hu, Y., Gerlt, J. A., Angerhofer, A., De Crécy-Lagard, V., & Bruner, S. D. (2021). Epoxyqueuosine Reductase QueH in the Biosynthetic Pathway to tRNA Queuosine Is a Unique Metalloenzyme. Biochemistry, 60(42), 3152-3161. https://doi.org/10.1021/acs.biochem.1c00164
Stack, T. M. M., & Gerlt, J. A. (2021). Discovery of novel pathways for carbohydrate metabolism. Current Opinion in Chemical Biology, 61, 63-70. https://doi.org/10.1016/j.cbpa.2020.09.005
Zallot, R., Oberg, N., & Gerlt, J. A. (2021). Discovery of new enzymatic functions and metabolic pathways using genomic enzymology web tools. Current Opinion in Biotechnology, 69, 77-90. https://doi.org/10.1016/j.copbio.2020.12.004
Garber, J. M., Nothaft, H., Pluvinage, B., Stahl, M., Bian, X., Porfirio, S., Enriquez, A., Butcher, J., Huang, H., Glushka, J., Line, E., Gerlt, J. A., Azadi, P., Stintzi, A., Boraston, A. B., & Szymanski, C. M. (2020). The gastrointestinal pathogen Campylobacter jejuni metabolizes sugars with potential help from commensal Bacteroides vulgatus. Communications biology, 3(1), . https://doi.org/10.1038/s42003-019-0727-5