Small molecules to manipulate peptide binding to MHC class I molecules, an optimized method for the generation of MHC tetramers by peptide exchange
Immunology
Cell Biology
Final Report Abstract
MHC class I molecules are transmembrane proteins that bind peptides (derived from proteolysis) inside the cell and carry them to the cell surface to present them to cytotoxic T cells. In this way, the immune system surveys the peptidome of every cell in the body. If inside a cancer cell or a virus-infected cell, a novel protein is made, its peptides are presented at the cell surface, and the T cells can induce the apoptosis of the preventing cell. This way, the spread of the cancer or the virus is inhibited. To find out whether in the body of a patient, T cells are present that react with peptides from the tumor, researchers use those peptides in complex with recombinant class I molecules, made into tetrameric form as so-called class I tetramers, to stain the patient's T cells in flow cytometry. In order to test many tumor peptides, many tetramers are required. Prior to our work, class I molecules could not be made in an empty form, and so, making many tetramers was tedious, slow, and expensive. We initially tried to optimize our previous method of peptide exchange catalyzed by small molecules to allow the generation of many tetramers. We realized that this was difficult to do, but at the same time, we discovered that those small molecules, dipeptides, allowed us to turn the disulfide-stabilized class I molecules that we had invented earlier into empty class I molecules. This is achieved by folding the disulfide mutants with the help of the dipeptides, and then rinsing out the dipeptides by gel filtration chromatography. The empty peptide binding site does not denature because it is held together by the disulfide bond. We found that the disulfide-stabilized empty class I molecules, once bound with peptide, are functional in the assay as described above just as wild type class I molecules. We believe that they can be used in the future to identify, characterize, isolate, and stimulate T cells, and that this will especially be useful in the immunotherapy of cancer and viral diseases. For example, in a therapy called adoptive transfer, T cells that recognize cancer cells are isolated from the blood of the patient, stimulated, multiplied, and returned to the patient to attack the cancer.
Publications
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Empty peptide-receptive MHC class I molecules for efficient detection of antigen-specific T cells Science Immunology 4 (2019)
Saini, Sunil Kumar; Tamhane, Tripti; Anjanappa, Raghavendra; Saikia, Ankur; Ramskov, Sofie; Donia, Marco; Svane, Inge Marie; Jakobsen, Søren Nyboe; Garcia-Alai, Maria; Zacharias, Martin; Meijers, Rob; Springer, Sebastian & Hadrup, Sine Reker
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High-throughput peptide-MHC complex generation and kinetic screenings of TCRs with peptidereceptive HLA-A*02:01 molecules. Science Immunology 4 (2019)
Moritz, Andreas; Anjanappa, Raghavendra; Wagner, Claudia; Bunk, Sebastian; Hofmann, Martin; Pszolla, Gabriele; Saikia, Ankur; Garcia-Alai, Maria; Meijers, Rob; Rammensee, Hans-Georg; Springer, Sebastian & Maurer, Dominik
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Successive crystal structure snapshots suggest the basis for MHC class I peptide loading and editing by tapasin. Proc. Natl. Acad. Sci. USA (2019)
Hafstrand, Ida; Sayitoglu, Ece Canan; Apavaloaei, Anca; Josey, Benjamin John; Sun, Renhua; Han, Xiao; Pellegrino, Sara; Ozkazanc, Didem; Potens, Renée; Janssen, Linda; Nilvebrant, Johan; Nygren, Per-Åke; Sandalova, Tatyana; Springer, Sebastian; Georgoudaki, Anna-Maria; Duru, Adil Doganay & Achour, Adnane
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Structures of peptide-free and partially loaded MHC class I molecules reveal mechanisms of peptide selection. Nature Communications 11, 1314 (2020)
Anjanappa, Raghavendra; Garcia-Alai, Maria; Kopicki, Janine-Denise; Lockhauserbäumer, Julia; Aboelmagd, Mohamed; Hinrichs, Janina; Nemtanu, Ioana Maria; Uetrecht, Charlotte; Zacharias, Martin; Springer, Sebastian & Meijers, Rob
