Stephen G. Withers

Profile Details

Position/Role

Director

Website

http://www.chibi.ubc.ca/faculty/withers/labhome

Research Interests

Coming from a background in the investigation of enzyme mechanisms, protein structure/function relationships, enzyme inhibitor design/synthesis and protein engineering we are increasingly adopting and developing tools to carry out such studies in a high-throughput manner. Our particular interest lies in enzymes that synthesise and degrade glycoconjugates such as glycolipids and glycoproteins and polysaccharides such as cellulose and starch.

One major area of interest is in the use of directed evolution approaches to engineer enzymes for the efficient synthesis of glycoconjugates, both by evolving natural glycosyl transferases and through the engineering of Glycosynthases, which are mutant glycosidases developed by our laboratory. To that end we are developing high-throughput screens that allow more than a million transformed bacteria to be scanned per hour. Enzymes so developed have now been licensed by industry for commercial syntheses.

Another area of interest is in the discovery of enzyme inhibitors or chaperones as novel pharmaceuticals through high-throughput screening. Recent successes include potential treatments for diabetes and Tay Sachs disease, with work ongoing on therapies for influenza and Chagas disease.

A third area of interest is in the development of new, targeted proteomics reagents to probe the presence and identities of specific families of enzymes, and we have recently published the first quantitative measures of this type using ICAT-like reagents. Given our deep experience with the enzymes involved in cellulose degradation we are applying these probes to better understanding of cellulosic biomass conversion.

Selected Publications

Glycobiology

Rich, J. R. and Withers, S. G. "Emerging methods for the production of homogeneous human glycoproteins" (2009) Nature Chemical Biology 5, 206-15.

Rao, F. V., Rich, J. R., Rakic, B., Buddai, S., Schwartz, M., Johnson, K., Bowe, C., DeFrees, S, Wakarchuk, W. W., Withers, S. G., Strynadka, N. C. J. "Structural insight into mammalian sialyltransferases" (2009) Nature Structural and Molecular Biology 16, 1186-8.

Engineering and Evolution

Hancock, S. M., Rich J. R., Caines, M. E. C., Strynadka, N. C. J. and Withers, S. G. "Designer enzymes for glycosphingolipid synthesis by directed evolution" (2009) Nature Chemical Biology 5, 508-514.

Yang, G and Withers, S. G. "Ultrahigh-throughput FACS-based screening for directed enzyme evolution" (2009) ChemBiochem 10, 2704–15.

Aharoni, A. and Withers, S. G. "Screening methodologies for glycosidic bond formation in the Protein Engineering Handbook" (2008) Lutz and Bornscheuer, Editors, John Wiley.

Hancock, S, Tarling, C, and Withers S.G. "High-throughput screening of cell lysates for ganglioside synthesis" (2008) Anal. Biochem. 382, 48-54.

Lairson, L. L., Watts, A. G., Wakarchuk, W. W. & Withers, S. G. “Glycosyl transferase substrate engineering: harnessing substrate promiscuity and expanding biological catalysis through transient chemical modification” (2006) Nature Chemical Biology 2, 724-8.

Aharoni, A., Thieme, K., Chiu, C. P.C., Buchini, S., Lairson, L. L., Chen, H., Strynadka, N.C.J., Wakarchuk, W.W. and Withers, S.G. “High throughput screening of glycosyltransferase libraries: Evolution of a sialyltransferase with a novel substrate specificity.” (2006) Nature Methods 3, 609-14.

Vaughan, M. D., Johnson, K., DeFrees, S., Tang, X., Warren, R. A. J., Withers, S. G. “Glycosynthase-mediated synthesis of glycosphingolipids” (2006) J. Am. Chem. Soc. 128, 6300-6301.

High-throughput screening for inhibitors

Kuntz, D. A., Tarling, C. A, Stick, R. V., Withers, S. G. and Rose, D. R. "Structural analysis of novel Golgi a-mannosidase II inhibitors from a focused glycosidase inhibitor screen" (2008) Biochemistry 47, 10058-68.

Tarling, C. A., Woods, K., Brastianos, H. C., Zhang, R., Brayer, G. D., Andersen, R. J. and Withers, S. G. “The search for novel human pancreatic α-amylase inhibitors: high-throughput screening of terrestrial and marine natural product extracts” (2008) ChemBiochem. 9, 433-8.

Tropak, M.B., Blanchard, J. E., Brown, E. D. and Mahuran, D. “High throughput screening for human lysosomal β-N-acetyl hexosaminidase inhibitors acting as pharmacological chaperones” (2007) Chem. & Biol. 14, 153-64.

Proteomic approaches

Hekmat, O., He, S., Warren, R. A. J. and Withers, S. G. “A mechanism-based ICAT strategy for comparing relative expression and activity levels of glycosidases in biological systems” (2008) J. Prot. Res. 7, 3282-92.

Hekmat, O., Florizone, C., Kim, Y-W., Eltis, L. D., Warren, R. A. J. and Withers, S. G. “Specificity fingerprinting of retaining α-1,4-glycanases in Cellulomonas fimi secretome using two fluorescent mechanism-based probes” (2007) ChemBiochem 8, 2125-32.

Williams, S. J., Hekmat, O. and Withers, S. G. “Synthesis and testing of mechanism-based protein profiling probes for retaining endo-glycosidases” (2006) ChemBiochem 7, 116-24.

Hekmat, O, Kim, Y-W, Williams, S. J., He, S. and Withers, S. G. “Active-site peptide ‘fingerprinting’ of glycosidases in complex mixtures by mass spectrometry: Discovery of a novel retaining β-1,4-glycanase in Cellulomonas fimi” (2005) J. Biol. Chem. 280, 35126-35.

History

Member for

1 year 26 weeks