Professor Research Research Areas: Organic Chemistry Biological Chemistry Research Interests: Enzymes are remarkable biocatalysts, not only for the dramatic rate accelerations (up to 1020 fold) that they provide, but also for the high degree of substrate specificity, regiospecificity and stereospecificity that these reactions exhibit. The work in my laboratory is focused on the chemical basis for how enzymes achieve such high rates and reaction specificity. Two groups of enzymes are currently under study in the laboratory: 1) Pyridoxal 5'-phosphate (PLP; vitamin B6) dependent enzymes, and, 2) Alcohol dehydrogenases. Tyrosine phenol-lyase and tryptophan indole-lyase are two PLP-dependent enzymes that catalyze the hydrolytic cleavage of tyrosine or tryptophan to phenol or indole, respectively, and ammonium pyruvate. Although the amino acid sequences and three dimensional structures of the two enzymes are very similar, these enzyme are specific for their physiological substrates. We are determining the chemical mechanisms of both enzymes by synthesis of substrate and transition state analogs, steady state and rapid-scanning stopped-flow kinetics, and by using site-directed mutagenesis. We are also altering the substrate specificity by mutagenesis to identify the amino acids which determine the reaction specificity. Another PLP-dependent enzyme being studied in my laboratory is kynureninase. We have cloned this enzyme from Pseudomonas fluorescens and Homo sapiens and we have studied the mechanism by steady state and pre-steady state kinetic methods. Recently, we have determined the crystal structures of bacterial and human kynureninases, and we are determining the structural basis for the differences in reaction specificity. We have also synthesized potent mechanism based inhibitors of kynureninase that could be useful as drugs. In other work, we are studying a thermostable secondary alcohol dehydrogenase (SADH) isolated from a thermophilic bacterium. We demonstrated a novel temperature dependent reversal of stereospecificity of SADH in the reaction of 2-butanol. We are currently investigating a mutant SADH with specificity for aromatic substrates. A new project uses hydrostatic pressure as a probe of conformational changes in enzymes and proteins. (This research was partially supported by a grant from the National Institutes of Health and the National Science Foundation.) Selected Publications Selected Publications: 2019-2026 Drago, V. N., Blakeley, M. P., Phillips, R. S., & Kovalevsky, A. (2026). Neutron diffraction reveals protonation states in pyridoxal‐5′‐phosphate‐free and glycine external aldimine‐bound serine hydroxymethyltransferase. The FEBS Journal, 293(2), 582-597. Phillips, R. S., Tran, X., & Mangano, B. (2025). Ornithine racemase uses a catalytic cysteine. Biochimica et Biophysica Acta (BBA)-Proteins and Proteomics, 141120. Baine, J. M., Duhoo, Y., Doukov, T., Desfosses, A., Bisello, G., Beio, M. L., Bauer, O., Perduca, M., Bacia-Verloop, M., Bertoldi, M., Phillips, R.S. & Berkowitz, D. B. (2025). α-Hydrazino Acids Inhibit Pyridoxal Phosphate-Dependent Decarboxylases via “Catalytically Correct” Ketoenamine Tautomers: A Special Motif for Chemical Biology and Drug Discovery?. ACS catalysis, 15(10), 8204-8218. Ma, W., McBride, J. T., & Phillips, R. S. (2025). Nitration of deactivated aromatic compounds using Lithium nitrate. Tetrahedron Letters, 159, 155515. Phillips, R. S., Brown, S. M., & Patel, R. S. (2024). Structural snapshots of Proteus vulgaris tryptophan indole-lyase reveal insights into the catalytic mechanism. ACS catalysis, 14(15), 11498-11511. Drago, V. N., Phillips, R. S., & Kovalevsky, A. (2024). Universality of critical active site glutamate as an acid–base catalyst in serine hydroxymethyltransferase function. Chemical Science, 15(32), 12827-12844. Phillips, R. S., & Hoang, K. N. N. (2022). The Y430F mutant of Salmonella d-ornithine/d-lysine decarboxylase has altered stereospecificity and a putrescine allosteric activation site. Archives of biochemistry and biophysics, 731, 109429. Dinh, T., Rahn, K. T., & Phillips, R. S. (2022). Crystallographic snapshots of ternary complexes of thermophilic secondary alcohol dehydrogenase from Thermoanaerobacter pseudoethanolicus reveal the dynamics of ligand exchange and the proton relay network. Proteins: Structure, Function, and Bioinformatics, 90(8), 1570-1583. Phillips, R. S., Jones, B., & Nash, S. (2022). M379A Mutant Tyrosine Phenol‐lyase from Citrobacter freundii Has Altered Conformational Dynamics. ChemBioChem, 23(13), e202200028. Li, Jun, Tung Dinh, and Robert Phillips. "The crystal structure of the S154Y mutant carbonyl reductase from Leifsonia xyli explains enhanced activity for 3, 5-bis (trifluoromethyl) acetophenone reduction." Archives of Biochemistry and Biophysics 720 (2022): 109158. Phillips, Robert S., Samuel C-K. Ting, and Kaitlin Anderson. "Structure and mechanism of d-Glucosaminate-6-phosphate ammonia-lyase: a novel octameric assembly for a pyridoxal 5′-phosphate-dependent enzyme, and unprecedented stereochemical inversion in the elimination reaction of a D-amino acid." Biochemistry 60.20 (2021): 1609-1618. Phillips, Robert S., and Austin P. Harris. "Structural basis of the stereochemistry of inhibition of tryptophan synthase by tryptophan and derivatives." Biochemistry 60.3 (2021): 231-244. Phillips, R. S., Craig, S., Kovalevsky, A., Gerlits, O., Weiss, K., Iorgu, A. I., Heyes, D.J. & Hay, S. (2019). Pressure and temperature effects on the formation of aminoacrylate intermediates of tyrosine phenol-lyase demonstrate reaction dynamics. ACS Catalysis, 10(3), 1692-1703. Phillips, R. S., Poteh, P., Krajcovic, D., Miller, K. A., & Hoover, T. R. (2019). Crystal structure of d-ornithine/d-lysine decarboxylase, a stereoinverting decarboxylase: implications for substrate specificity and stereospecificity of fold III decarboxylases. Biochemistry, 58(8), 1038-1042.
