Geometric factors influencing hydrogen-deuterium exchange: Detection of mobile water molecules in the peptide-MHCI - binding interface (LB291)

Hydrogen deuterium exchange (HDX) has long been used as an important technique to study conformational changes of proteins in solution, protein folding as well as protein ligand interactions. HDX is thought to detect exposure of protein backbone amide (bNH) hydrogens to solvent or internal water. However, it is not known how this exposure is defined in geometric terms, especially for internal water-filled cavities. The experiments described here was designed to gain a better understanding of the influence of factors like distance and angle of orientation of bNH-groups relative to internal water pockets. We used Major Histocompatibility Class I (MHCI)-peptide complexes as a model to address this question. We expressed MHCI heavy and light chains and reconstituted MHCI-peptide complexes with synthetic immunogenic peptides. We measured HDX using LC-MS, and characterized structural parameters of water-filled pockets with the help of computer programs like Pymol and Surfnet. We have successfully developed a mo...