title: Studies on calcium ion-induced conformation changes in the actin-tropomyosin-troponin system by fluorimetry. III. Changes in the conformation of tropomyosin associated with functional states.
abstract: The local conformational changes in the tropomyosin molecule under various conditions were studied by means of fluorimetry using SH-directed fluorescent dyes, N-(1-anilinonaphthyl-4)maleimide (ANM) and N-(3-pyrene)maleimide (PRM). 1. The fluorescence intensity, polarization and the emmission maximum of ANM-tropomyosin were found to be susceptible to ionic strength, but in different ways. The changes in these parameters suggest that the fluorescence-labeled sulfhydryl group or groups become more buried in a hydrophobic internal region by salt-induced depolymerization of aggregate and by adding F-actin to tropomyosin. 2. Titration of the labeled tropomyosin with F-actin revealed a cooperative nature in ANM labeling and a simple saturation kinetics in PRM labeling. The dissociation constant of F-actin to PRM-tropomyosin was calculated to be 5.8-10(-6) M. 3. Temperature dependence of the fluorescence polarization showed a thermal transition in the conformation of ANM- or PRM-tropomyosin at around 30 degrees C. Flexibility or segmental motion of the region containing the fluorophore was suppressed significantly on adding troponin and markedly on adding F-actin. 4. Measurements of the quantum yield and polarization of the ANM-tropomyosin-F-actin complex suggested that troponin strengthened the binding between the two proteins and that Ca2+ reversed this effect.
mesh_terms: D000199:Actins; D001665:Binding Sites; D002118:Calcium; D046911:Macromolecular Substances; D008301:Maleimides; D009124:Muscle Proteins; D009281:Naphthalenes; D011485:Protein Binding; D011487:Protein Conformation; D011789:Quantum Theory; D013050:Spectrometry, Fluorescence; D014335:Tropomyosin; D014336:Troponin
chemical_list: D000199:Actins; D046911:Macromolecular Substances; D008301:Maleimides; D009124:Muscle Proteins; D009281:Naphthalenes; D014335:Tropomyosin; D014336:Troponin; D002118:Calcium