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The immune system is more than antibodies, T-cells and MHC molecules. For instance, the innate branch of the immune system includes a type of cells known as natural killer (NK) cells that can survey other cells for stress markers produced when that cell is infected by a virus or transformed into a cancer cell. Once one of these markers is detected, the NK cell will destroy the stressed cell, thus stopping the spread of an infection or tumor. One of the most interesting receptors on the NK cell surface that mediates these interactions is called NKG2D. The McFarland Lab studies the interactions of NKG2D with its various protein ligands, including MIC-A and MIC-B. Study of these interactions will lead to insights that could impact our understanding of how protein-protein interfaces work and could impact cancer therapies involving NK cells.

We are currently investigating two areas of inquiry involving structural investigations of NKG2D-ligand interfaces. We began with the observation that the crystal structure of unbound MIC-A was lacking a region of disordered structure in the region of the a2 helix, squarely in the middle of the NKG2D binding footprint. Electron density for this region was observed in the NKG2D—MIC-A complex crystal structure in the expected helical conformation. A computer program, RosettaDesign, was used to redesign eight residues at the “interface” between the MIC-A a2 helix and the MIC-A core in order to pre-order this loop, stabilize the MIC-A surface and affect NKG2D binding kinetics and/or thermodynamics. Several potential stabilizing designs were ranked by RosettaDesign, and we are in the process of mutating MIC-A and producing mutant proteins. Many of the proteins so far show signs of being structurally stabilized relative to wild-type MIC-A. MIC-B also shows signs of disorder in the a2 region in the unbound form, and it is being prepared for analysis and mutation. If these processes result in a higher-affinity MIC protein, such a reagent could have laboratory or therapeutic use.

We are also investigating the covalent modification of positively charged residues at NKG2D ligands’ surfaces with dihydroxyacetone (DHA, commonly used in sunless tanning lotions). Different ligands are inactivated by DHA to different extents, and studies are underway to understand this modification at a structural level and increase our knowledge of these protein-protein interfaces.