Fluorescence Approaches for the Characterization of the Peripheral Membrane Binding of Protein Applied for the Blood Coagulation Protein Prothrombin

摘要

Different fluorescence spectroscopic approaches for the characterization of the peripheral membrane binding of proteins are summarized and compared for the blood coagulation protein prothrombin. Basically, the problem of membrane binding can be approached from both the protein side, using either intrinsic tryptophan fluorescence or dye-labeled protein, as typically employed in fluorescence recovery after photobleaching experiments, or from the membrane aide, studying solvent relaxation, fluorescence anisotropy or excimer formation of suitable membrane probes. For the case of prothrombin and its fragment 1 the combined use of these fluorescence methods shows a slightly tighter binding of the fragment 1 portion in the case of the entire prothrombin molecule compared to the isolated fragment 1 while no evidence could be obtained for hydrophobic membrane binding sites in the "nonfragment 1" part of prothrombin. Tryptophan fluorescence of fragment 1 is able to detect differences in binding in response to the kind of procoagulant membrane lipid.