Scientific Report 2008
Bioorganic and Synthetic Chemistry
C.-H. Wong, C. Bennett, S. Dean, S. Ficht,
Y. Fu, W. Greenberg, R. Guy, S. Hanson, Z. Hong, D.-R. Hwang, M. Imamura, K.
Kishikawa, J.-C. Lee, P.-H. Liang, L. Liu, T. Polat, S.-K. Wang, Y.-Y. Yang
We develop new
chemical and enzymatic strategies for synthesis of bioactive small molecules and
biomolecules. We use the methods to probe carbohydrate-mediated recognition events
important in cancer, bacterial infections, and viral infections, including HIV disease
We have developed new methods for sugar-assisted
ligation of glycopeptides for synthesis of homogenous glycoproteins. We have used
the methods in conjunction with enzymatic glycosylation techniques to assemble complex
glycopeptides by chemical synthesis, and we are optimizing the techniques to achieve
the total synthesis of therapeutic glycoproteins. Glycoproteins are expressed in
vivo as complex mixtures of glycoforms, a situation that hinders efforts to study
the role of glycosylation in protein folding, stability, and function. By synthesizing
pure glycoforms, we can characterize in molecular detail the effects of glycans
on protein function.
Using chemical techniques such as programmable
1-pot oligosaccharide synthesis, as well as enzymatic synthesis, we create glycoarrays
on glass slides for high-throughput quantitative analysis of protein-carbohydrate
interactions. These arrays are being used to study the binding specificity of carbohydrate-binding
receptors and antibodies. We have applied aldolases, glycosyltransferases, glycosidases,
and other enzymes to develop practical new methods of synthesizing molecules such
as iminocyclitols, which are inhibitors of glycosidases and other enzymes, and glycopeptides,
and other glycoconjugates. Using directed evolution, we are evolving these enzymes
to catalyze new reactions and synthesize new molecules of pharmaceutical relevance.
We are using our synthetic methods to
discover inhibitors and therapeutic agents in several diseases related to carbohydrates.
Current targets include bacterial transglycosidase, sulfatases, and glycoprocessing
enzymes involved in the biosynthesis of carbohydrates that mediate cancer metastasis,
inflammation, and viral infection. Enzymatically synthesized iminocyclitols are
being investigated as treatments for osteoarthritis and Gaucher disease. Inspired
by the broadly neutralizing anti-HIV antibody 2G12, which recognizes a dense array
of oligomannose displayed on HIV gp120, we are designing dendrimeric oligomannose
structures for development of an HIV vaccine. In collaboration with D.R. Burton,
Department of Immunology, we are testing the immunogenicity of these constructs.
We have designed glycolipid ligands for CD1, which activate natural killer T cells
and are a promising new immunotherapeutic approach for treatment of bacterial and
viral infections and cancer. They may also be useful as adjuvants in vaccine development.
Glycoproteomics And Molecular Glycobiology
Using metabolic oligosaccharide engineering,
we have developed methods for incorporating tagged sugars into glycans expressed
on mammalian cells. The engineered glycans can be labeled with a variety of molecules
by using click chemistry. One application is glycan-specific fluorescent labeling,
which is used for fluorescent imaging to compare glycosylation patterns of different
cells, such as normal vs cancer cells or cancer cells vs cancer stem cells. We found
that protein fucosylation and sialylation are both elevated in cancer cell lines.
A second application of this chemistry
is GIDmap, a new method for glycoproteomic analysis (Fig. 1). Whole cells are fed
with tagged sugars, and after biochemical incorporation of the sugars into cellular
glycoproteins, click chemistry is used to attach a handle for purification of tagged
proteins. Mass spectrometric proteomic methods are then used to identify proteins
that are differentially glycosylated. We are using GIDmap to identify proteins that
are aberrantly glycosylated in different stages of cancer. These cancer-associated
glycoproteins may be useful as biomarkers for diagnostics or as targets for therapeutic
1. GIDmap glycoproteomic analysis via metabolic oligosaccharide engineering.
Bennett, C.S., Dean, S.M., Payne,
R.J., Ficht, S., Brik, A., Wong, C.-H.
Sugar-assisted glycopeptide ligation with complex oligosaccharides: scope and limitations.
J. Am. Chem. Soc. 130:11945, 2008.
Ficht, S., Payne, R.J., Guy, R.T.,
Wong, C.-H. Solid-phase synthesis
of peptide and glycopeptide thioesters through side-chain-anchoring strategies.
Chem. Eur. J. 14:3620, 2008.
Giffin, M.J., Heaslet, H., Brik, A.,
Lin, Y.-C., Cauvi, G., Wong, C.-H., McRee, D.E., Elder, J.H., Stout, C.D., Torbett,
B.E. A copper(I)-catalyzed
1,2,3-triazole azide-alkyne click compound is a potent inhibitor of a multidrug-resistant
HIV-1 protease variant. J. Med. Chem. 51:6263, 2008.
Hanson, S.R., Greenberg, W.A., Wong
C.-H. Probing glycans with
the copper(I)- catalyzed [3+2] azide-alkyne cycloaddition. QSAR Comb. Sci. 26:1243,
Kinjo, Y., Pei, B., Bufali, S., Raju,
R., Richardson, S.K., Imamura, M., Fujio, M., Wu, D., Khurana, A., Kawahara, K.,
Wong, C.-H., Howell, A.R., Seeberger, P.H., Kronenberg, M.
Natural Sphingomonas glycolipids vary greatly in their ability to activate
natural killer T cells. Chem. Biol. 15:654, 2008.
Liang, P.-H., Imamura, M., Li, X.,
Wu, D., Fujio, M., Guy, R., Wu, B.-C., Tsuji, M., Wong, C.-H .
Quantitative microarray analysis of intact glycolipid-CD1d interaction and correlation
with cell-based cytokine production. J. Am. Chem. Soc. 130:12348, 2008.
Liang, P.-H., Wu, C.-Y., Greenberg,
W.A., Wong, C.-H. Glycan arrays: biological and medical applications. Curr. Opin. Chem. Biol. 12:86, 2008.
Northen, T.R., Lee, J.-C., Hoang,
L., Raymond, J., Hwang, D.-R., Yannone, S.M., Wong, C.-H., Siuzdak, G. A
nanostructure-initiator mass spectrometry-based enzyme activity assay. Proc. Natl.
Acad. Sci. U. S. A. 105:3678, 2008.
Payne, R.J., Ficht, S., Greenberg, W.A., Wong, C.-H. Cysteine-free
peptide and glycopeptide ligation by direct aminolysis. Angew. Chem. Int. Ed. 47:4411,
Sugiyama, M., Hong, Z., Liang, P.-H.,
Whalen, L.J., Greenberg, W.A., Wong, C.-H. D-Fructose-6-phosphate
aldolase-catalyzed one-pot synthesis of iminocyclitols. J. Am. Chem. Soc. 129:14811,
Wang, S.-K., Liang,
P.-H., Astronomo, R.D., Hsu, T.-L., Hsieh, S.-L., Burton, D.R., Wong, C.-H.
Targeting the carbohydrates on HIV-1: interaction of oligomannose dendrons with
human monoclonal antibody 2G12 and DC-SIGN. Proc. Natl. Acad. Sci. U. S. A. 105:3690,
Whalen, L.J., Greenberg, W.A., Mitchell,
M.L., Wong, C.-H. Iminosugar-based
glycosyltransferase inhibitors. In: Iminosugars: From Synthesis to Therapeutic
Applications. Compain, P., Martin, O.R. (Eds.). Wiley-VCH, Hoboken, NJ, 2007, p.
Wu, D., Fujio, M., Wong, C.-H.
Glycolipids as immunostimulating agents. Bioorg. Med. Chem. 16:1073, 2008.