David S. Waugh, Ph.D.
The Protein Engineering Section combines X-ray crystallography with strong molecular biology support to enhance and accelerate studies of protein function and drug development. A number of highly collaborative projects are currently underway, involving kinases, phosphatases, proteases and other enzymes. We also contribute to the development of new and improved methods and reagents for the production of recombinant proteins. In particular, we are interested in understanding how certain highly soluble proteins are able to prevent the aggregation and promote the proper folding of their fusion partners, and in exploiting proteases as reagents for the removal of affinity tags.
The Protein Engineering Section combines X-ray crystallography with strong molecular biology support to enhance and accelerate studies of protein function and drug development. A number of highly collaborative projects are currently underway.
With Dr. John (Jay) Schneekloth (CCR's Chemical Biology Laboratory), we are attempting to develop inhibitors/chemical probes of Ubc9, the lone SUMO-conjugating enzyme in humans. SUMOylation regulates a wide variety of biological processes, and defects in SUMO homeostasis are associated with many diseases including cancer. High-resolution crystal structures of Ubc9 with drug-like ligands are being sought and obtained by fragment screening and other complementary approaches.
Human tyrosyl-DNA phosphodiesterase 1 (Tdp-1) is the focus of another project. Tdp1 inhibitors are expected to act synergistically with topoisomerase 1-targeting drugs (camptothecins, indenoisoquinolines), topoisomerase 2 inhibitors (etoposide, doxorubicin), bleomycin, and DNA alkylating agents that are already used for cancer chemotherapy. In collaboration with Dr. Yves Pommier (CCR's Developmental Therapeutics Branch), we are seeking to elucidate the structural basis for inhibition of Tdp-1 by various compounds and, by extension, gain insight into how these inhibitors might be improved by rational design.
Protein phosphorylation is a reversible event that is carefully controlled by the opposing activities of kinases and phosphatases. Abnormal changes to this equilibrium often lead to deleterious effects such as deregulated cell growth and cancer. We are collaborating with Dr. Terrence Burke Jr. (CCR's Chemical Biology Laboratory), an expert in the field of protein tyrosine phosphatase (PTPase) inhibition, to develop inhibitors of PTPase epsilon, a membrane-bound enzyme that is up-regulated in human breast cancer cells and which is also a negative regulator of insulin receptor signaling. To facilitate this goal, we have recently determined a high-resolution crystal structure of the membrane-proximal catalytic domain.
Raf kinase is a central signaling molecule in the mitogen-activated protein kinase cascade that is constitutively activated by mutation at high frequency in certain cancers. Dimerization of the Raf kinase domain is an obligatory step in its activation. Crystal structures of dimeric Raf kinase in complex with a variety of ligands have been determined. However, the structure of monomeric Raf remains unknown, and hence the conformational changes that accompany dimerization and activation are poorly understood. Seeking to gain insight into this process, in collaboration with Dr. Deborah Morrison (CCR's Laboratory of Cell and Developmental Signaling), we are attempting to crystallize and determine the structure of a Raf kinase mutant (R481H) that is unable to dimerize.
Extramural collaborators include investigators at the United States Army Medical Institute of Infectious Diseases (Drs. Robert Ulrich and Kamal Saikh), with whom we are studying the role of the adaptor molecule MyD88 in the innate immune response and structure/function relationships in dual-specificity phosphatases. Working with Dr. Pamela Glass (USAMRIID) and Dr. Patricia Legler of the US Naval Research Laboratory, we are using X-ray crystallography to facilitate the development of inhibitors of Venezuelan Equine Encephalitis Virus (VEEV) protease and related enzymes from Chikungunya virus and Middle East Respiratory Syndrome coronavirus. To this end, a high-resolution crystal structure of VEEV protease has recently been determined.
Finally, the Protein Engineering Section continues to contribute to the development of new and improved methods and reagents for the production of recombinant proteins. In particular, we are interested in understanding how certain highly soluble proteins are able to prevent the aggregation and promote the proper folding of their fusion partners, and in exploiting proteases as reagents for the removal of affinity tags.
Selected Key Publications
Escherichia coli maltose-binding protein is uncommonly effective at promoting the solubility of polypeptides to which it is fused.Protein Sci. 8: 1668-74, 1999. [ Journal Article ]
Tobacco etch virus protease: mechanism of autolysis and rational design of stable mutants with wild-type catalytic proficiency.Protein Eng. 14: 993-1000, 2001. [ Journal Article ]
Three-dimensional structure of a macromolecular assembly that regulates type III secretion in Yersinia pestis.J Mol Biol. 346: 1147-61, 2005. [ Journal Article ]
Crystal structure of checkpoint kinase 2 in complex with NSC 109555, a potent and selective inhibitor.Protein Sci. 18: 92-100, 2009. [ Journal Article ]
Utilization of nitrophenylphosphates and oxime-based ligation for the development of nanomolar affinity inhibitors of the Yersinia pestis outer protein H (YopH) phosphatase.J. Med. Chem. 54: 2933-43, 2011. [ Journal Article ]
Dr. Waugh graduated from Yale University in 1982 with a B.S. in Molecular Biophysics & Biochemistry. He earned his Ph.D. in Biochemistry & Molecular Biology from Indiana University in 1989 under the direction of Dr. Norman Pace. After two years as a Postdoctoral Fellow in Dr. Robert Sauer's laboratory at the Massachusetts Institute of Technology, he became Director of the Macromolecular Engineering Laboratory at Hoffmann-La Roche. In 1996, Dr. Waugh established the Protein Engineering Section of the ABL Basic Research Program at NCI-Frederick. He moved to the NCI Center for Cancer Research in 1999 and gained tenure as a Senior Investigator in 2005.