Research and Teaching InterestsOur research focuses on the design of artificial model proteins that can perform a desired function. In one effort, we are developing novel enzymes that could catalyze specific non-biological transformation. In a parallel effort, the group is using protein engineering to design minimalist models of proteins involved in complex biological processes.
Novel membrane proteins for redox catalysis We have designed a stable, membrane soluble protein that utilizes a metal cofactor, heme, as active site. The design started from a natural membrane protein, Glycophorin A: by mutating two residues to histidine, we engineered a heme binding site in the hydrophobic pocket. The novel protein binds heme with sub-micromolar affinity and catalyzes the oxidation of several organic substrates; current efforts are directed to the creation of a library of mutants in order to optimize the catalytic activity. Possible applications of these model proteins are in environmental bioremediation and in the synthesis of fine chemicals. Methods utilized include computational modeling, solid phase synthesis, molecular biology techniques and enzymatic screening. A Novel Immunomodulatory Protein We are currently working on the development of mini-protein analogs of macrophage activating factor (Gc-MAF), a glycoprotein involved in the immune system activation. In Gc-MAF, a single glycosylation event on a threonine leads to the active form of the protein. Using computer visualization software, the site of glycosylation and the surrounding loop structure was identified and grafted on a smaller, stable scaffold. We prepared a prototype model by solid phase synthesis: we are currently testing this model for its ability to activate cultured macrophages. The experimental screening provides information used to improve the initial design. The ultimate goal in this project is to utilize the miniaturized proteins as pharmaceuticals in diseases such as acute inflammation and as adjuvant in cancer therapy. |
Publications
"Multivalent interactions with gp120 are required for the anti-HIV activity of cyanovirin.," Liu Y, Carroll JR, Holt LA, Mc Mahon J, Giomarelli B, Ghirlanda G., Biopolymers in press (2009)
"Modulation of protein stability by O-glycosylation in a designed Gc-MAF analog," Spiriti, J., Bogani F., van der Vaart A, and Ghirlanda, G., Biophysical Chemistry 134 157 (2008)
"Computational Biochemistry: Old Enzymes, New Tricks," G. Ghirlanda, Nature 453 164 (2008)
"Conformational gating of sugar binding to the antiviral protein cyanovirin revealed from the crystal structure at 1.35 Å resolution," Conformational gating of sugar binding to the antiviral protein cyanovirin revealed from the crystal structure at 1.35 Å resolution, Protein Science 17 939 (2008)
"Design of a functional membrane protein by engineering a heme-binding site in glycophorin A," J.M. Cordova, P.L. Noack, S.A. Hilcove, J.D. Lear, and G.Ghirlanda, J. Am. Chem. Soc 129 512-518 (2007)
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