| Popis: |
While molecular structures are available for many proteins, rarely is a structure determined in a protein's functional context. For proteins that bind with lipid membranes, traditional structural methods cannot probe the protein-membrane complex. X-ray reflectivity allows us to characterize the dynamic structure of these systems in physiologically relevant conditions. Given a structure of the protein, derived from techniques such as crystallography or nuclear magnetic resonance (NMR), it is possible to determine the structure of the protein-membrane complex using X-ray reflectivity. T-cell Immunoglobulin Mucin (TIM) family of proteins are involved in the recognition of phosphatidylserine (PtdSer), a cellular signal for apoptosis, in lipid membranes. The family has a conserved binding pocket that selectively recognizes PtdSer in the presence of calcium. While each member has varying binding sensitivity to the density of PtdSer in membranes, TIMs’ structural variations, as solved by crystallography and NMR, do not explain the variance in binding sensitivity. We have previously studied TIM4 and showed that peripheral interactions of particular residues on TIM4 with the membrane can explain TIM4's higher affinity for membranes with high densities of PtdSer. We have now extended the work to TIM1 and TIM3 in a similar effort to elucidate the structural basis of their binding sensitivities. Unlike TIM4, the crystal and NMR structures of TIM1 and TIM3, respectively, do not fit the X-ray reflectivity data well. We employ molecular dynamics (MD) to find the most likely conformations of TIM1 and TIM3 in complex with a lipid membrane and use the MD solved structures to fit X-ray reflectivity data. Using this combination of computational, experimental and statistical techniques, we are able to determine the most likely conformations of TIM1 and TIM3 in complex with a lipid film. |
