We are interested in structure-function relationships of enzymes and enzyme evolution i.e. the relationship between sequence/structural homology and function. Many of the enzymes we work with also form heteromeric complexes and are ideal candidates for investigating how quaternary structures and protein-protein interactions modulate the functions of biological catalysts.
Projects in our lab include
(1) Steroid degradation by Mycobacterium tuberculosis and related bacteria
The cholesterol degradation pathway in M. tuberculosis is important for the persistence of the bacteria in host macrophages, and it therefore represents a potential target for development of new antibiotics against the pathogen. We are studying a number of enzymes involved in the steroid side-chain degradation pathway. These enzymes are related to those involved in fatty acid beta-oxidtion but they possess unique quaternary structures that provide the molecular determinants for binding of the large steroid substrates.
(2) Lignin/ Aromatic Pollutants and toxins Degradation
We study enzymes involved in aromatic/cyclic hydrocarbon and toxin degradation. In the PCBs degradation pathway, the aldolase, BphI, is in complex with a dehydrogenase, BphJ, that catalyze the transformation of the aldehyde product from the aldolase reaction to acyl CoAs. Aldehyde products, of various lengths and subsitution, are channeled from the aldolase to the dehydrogenase via a molecular tunnel, ensuring that the toxic aldehydes do not poison the cells. We are investigating how vectorial movements of aldehydes through the enzyme complex are accomplished through protein dynamics using combination of biochemical and structural approaches.
(3) Detoxification of mycotoxins
In collaboration with Dr. Ting Zhou at Agriculture Agri-food Canada, we are isolating and characterizing enzymes capable of detoxifying the mycotoxins, deoxynivalenol and patulin. These mycotoxins contaminate grains and fruit juices.
Our laboratory is situated at the Science complex providing us with access to state-of-the-art facilities and instruments.
We employ a wide variety of interdisplinary techniques in our research, including
1. Molecular Genetics – PCR, Gene cloning, Gene knockout, Directed/Random mutagenesis
2. Protein Purification
3. Enzymology – Steady-state and Pre-steady-state kinetics
4. Spectroscopy – UV-Visible, Fluorescence, Circular Dichroism, Nuclear Magnetic Resonance (NMR)
5. Analytical Chemistry including HPLC and Mass spectrometry
6. X-ray crystallography
7. Molecular modelling
1. Abraham, N, Chan, E.T.S., Zhou, T. and Seah, S.Y.K. (2022) Microbial detoxification of mycotoxins in food. Frontiers in Microbiology (in-press)
2. Abraham, N, Schroeter, K.L., Zhu, Y., Chan, J., Evans, N., Kimber, M.S., Carere, J., Zhou, T. and Seah, S.Y.K. (2022) Structure-function characterization of an aldo-keto reductase involved in detoxification of the mycotoxin, deoxynivalenol. Scientific Reports 12:14737.
2. Schroeter, K.L., Abraham, N., Rolfe, N., Barnshaw, R., Diamond, J. and Seah, S.Y.K. (2022) Bacterial hydratases involved in steroid side-chain degradation have distinct substrate specificities. J. Bacteriol. 204:e0023622
3. Chan E.T.S., Zhu, Y., Li, X.Z., Zhou, T. and Seah S.Y.K. (2022) Characterization of two dehydrogenases from Gluconobacter oxydans Involved in the transformation of patulin to ascladiol. Toxins (Basel) 14:423
4. Stirling, A.J., Gilbert, S.E., Conner, M., Mallette, E., Kimber, M.S., Seah S.Y.K. (2020) A key glycine in bacterial steroid-degrading acyl-CoA dehydrogenases allows Flavin-ring repositioning and modulates substrate side chain specificity. Biochemistry 59:4081-4092
5. Aggett R., Mallette, E., Gilbert, S.E., Vachon, M.A., Schroeter, K.L., Kimber, M.S. and Seah S.Y.K. (2019) The steroid side chain cleaving aldolase Ltp2-ChsH2DUF35 is a thiolase superfamily member with a radically repurposed active site. J. Biol. Chem. 294:11934-11943.
6. Gilbert, S. Hood, L. and Seah, S.Y.K. (2017) Characterization of an aldolase involved in cholesterol side chain degradation in Mycobacterium tuberculosis. J. Bacteriol. 200: e00512-17
7. Mazurkewich S and Seah SY.K (2016) Investigation into the Mode of Phosphate Activation in the 4-Hydroxy-4-Methyl-2-Oxoglutarate/4-Carboxy-4-Hydroxy-2-Oxoadipate Aldolase from Pseudomonas putida F1. PLoS One. 14;11:e0164556.
8. Mazurkewich, S., Brott, A.S., Kimber, M.S. and Seah S.Y.K. (2016) Structural and Kinetic Characterization of the 4-Carboxy-2-Hydroxymuconate Hydratase from the Gallate and Protocatechuate 4,5-Cleavage Pathways of Pseudomonas putida KT2440 J. Biol. Chem. 291:7669-7686.
9. Ruprecht, A., Maddox, J., Stirling, A.J., Visaggio, N. and Seah S.Y.K. (2015) Characterization of novel Acyl-CoA dehydrogenases involved in bacterial steroid degradation. J. Bacteriol. 197:1360-1367
10. Mazurkewich, S., Wang, W. and Seah S.Y.K. (2014) Biochemical and structural analysis of RraA proteins to decipher their relationships with 4-hydroxy-4-methyl-2-oxoglutarate/4-carboxy-4-hydroxy-2-oxoadipate aldolases. Biochemistry 53:542-553
11. Carere, J., McKenna, S.E., Kimber, M.S. and Seah S.Y.K. (2013) Characterization of an Aldolase-Dehydrogenase Complex from the Cholesterol Degradation Pathway of Mycobacterium tuberculosis. Biochemistry 52:3502-3511.
12. Coincon, M., Wang, W., Sygusch, J., Seah S.Y.K. (2012) Crystal Structure of Reaction Intermediates in Pyruvate Class II Aldolase: Substrate cleavage, enolate stabilization, and substrate specificity. J Biol Chem. 287:36208-36221
13. Baker, P., Carere, J. and Seah, S.Y.K. (2012) Substrate Specificity, Substrate Channeling, and Allostery in BphJ: An Acylating Aldehyde Dehydrogenase Associated with the Pyruvate Aldolase BphI. Biochemistry 51:4558-4567.
14. Baker, P., Hillis, C., Carere, J. and Seah, S.Y.K. (2012) Protein-protein interactions and substrate channeling in orthologous and chimeric aldolase-dehydrogenase complexes. Biochemistry 51:1942-1952.
15. Baker, P and Seah, S.Y.K. (2012) Rational Design of Stereoselectivity in the Class II Pyruvate Aldolase BphI. J Am Chem Soc. 134:507-513.
16. Ng, F.S.W., Wright, D. and Seah, S.Y.K. (2011) Characterization of a phosphotriesterase-like lactonase from Sulfolobus solfataricus and its immobilization for quorum quenching. Appl Environ Microbiol 77:1181-1186