| Membrane proteins have been a challenge to study since they are soluble only in the presence of detergents and it has been difficult to obtain very high resolution crystal structures. Although there are several thousand soluble protein structures in the protein database, only about 100 membrane protein structures are accumulated. However the highest resolution yet is still well over 1 A, which is required to observe protons. We are interested in the mechanism of proton transfer within several membrane proteins, and are using FTIR spectroscopy to obtain information about this.;We are studying cytochrome c oxidase from R. sphaeroides, the photosynthetic reaction center from R. sphaeroides , and bacteriorhodopsin from H. salinarum, which are all membrane proteins that transfer protons partially or entirely across the cell membrane as part of their function. Proton movements inside the protein are achieved by "hopping" along hydrogen bonds involving either water molecules, polar residues or protonatable residues. Cytochrome c oxidase catalyzes the reduction of molecular oxygen coupled to proton pumping. In order to understand how the proton pump works, in cytochrome c oxidase, it is critical to know the timing of the various protonation/deprotonation reactions during the catalytic cycle.;Fourier Transform Infrared Spectrsocopy (FTIR) is a powerful technique that is well suited to the investigation of protonation/deprotonation reactions of individual side chains within proteins, as well as any changes that might occur in the strength of H-bonds any change in the overall protein structure. The method can be applied in a static manner, revealing differences between two defined states of a protein, or can be done in a time-dependent manner in the microsecond/millisecond time range. |
