Research Focus

Molecular Bioengineering, Protein Engineering, Therapeutic Protein Biotechnology.

Engineers now have the tools to design biological products and processes at the molecular level. Proteins are of particular therapeutic interest, because proteins mediate most biochemical processes both inside and outside cells. The ability to manipulate the strength and specificity of protein binding events provides tremendous leverage for the development of novel biopharmaceuticals. We are developing powerful new tools for protein engineering, and applying them both to particular disease targets and to bettering our understanding of protein structure/function relationships. In the absence of predictive capabilities for protein design, a directed evolution or combinatorial library screening strategy can be effectively applied to alter protein properties in a desired fashion. We apply quantitative engineering analyses of the relevant kinetic and statistical processes to develop optimal search strategies on the protein ìfitness landscape.î In particular, we have developed a method for protein display on the surface of yeast cells that, for example, enabled engineering of a noncovalent protein-ligand bond with a dissociation half-time over one week. We are engineering potential protein biopharmaceuticals in areas where molecular understanding of disease pathology is sufficient to hypothesize particular objective functions to target. For example, antibodies can be used to target cell-killing modalities to cancerous cells, given sufficiently strong and specific binding properties. Growth factors that carry signals between cells do so via particular binding events that, if manipulated to alter intracellular trafficking or signalling outcomes, could alter immune responses in precisely defined ways. Finally, viral and nonviral vectors for gene therapy could be targeted to specific cells and tissues via alteration of an exchangeable antibody recognition module. Altered proteins developed in this work can also provide a potential vehicle for new insights into the mechanisms of protein-ligand binding. We are performing biophysical analyses of the kinetic, thermodynamic, and structural aspects of engineered protein function in order to contribute to an improved understanding of protein binding processes.

Selected publications

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Graff CP, Wittrup KD. Theoretical analysis of antibody targeting of tumor spheroids: importance of dosage for penetration, and affinity for retention. Cancer Res. 2003 Mar 15;63(6):1288-96.

Feldhaus MJ, Siegel RW, Opresko LK, Coleman JR, Feldhaus JM, Yeung YA, Cochran JR, Heinzelman P, Colby D, Swers J, Graff C, Wiley HS, Wittrup KD.
Flow-cytometric isolation of human antibodies from a nonimmune Saccharomyces cerevisiae surface display library. Nat Biotechnol. 2003 Feb;21(2):163-70.

Wittrup KD. Protein engineering by cell-surface display. Curr Opin Biotechnol. 2001 Aug;12(4):395-9. Review.

Yeung YA, Wittrup KD. Quantitative screening of yeast surface-displayed polypeptide libraries by magnetic bead capture. Biotechnol Prog. 2002 Mar-Apr;18(2):212-20.

Boder ET, Midelfort KS, Wittrup KD. Directed evolution of antibody fragments with monovalent femtomolar antigen-binding affinity.
Proc Natl Acad Sci U S A. 2000 Sep 26;97(20):10701-5.

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