title: Interaction of actinogelin with actin. No nucleation but high gelation activity.
abstract: Nucleation activity of actin polymerization of actinogelin, a calcium-sensitive F-actin cross-linking protein from rat liver, was measured by a fluorescence enhancement method using pyrenyl-actin and by high shear viscometry. No stimulation of nucleation by the addition of actinogelin was observed under several ionic conditions using the fluorescent method. Similar results were also obtained by viscometry. Therefore, it can be concluded that actinogelin has no nucleation activity for actin polymerization. By electron microscopy, it was found that actinogelin molecule has a dumbbell shape, binds to side of F-actin through its end(s), and cross-links actin filaments by binding with its two ends. It was also found that meshwork formation occurred in low Ca2+ conditions from F-actin and actinogelin. Under non-gelling high Ca2+ conditions, binding of actinogelin along the side of F-actin with its one end was still detected in accordance with the binding assay using ultracentrifugation and protein determination. Under low Ca2+ conditions, the critical gelling concentration of actinogelin measured by low shear viscometry at 20 degrees C was 6 micrograms/ml for 250 micrograms/ml of actin. Comparing this value with those of the other actin cross-linking proteins, it was found that actinogelin was one of proteins with the highest gelation activity. On the other hand, gelation activity of actinogelin in high Ca2+ conditions was one order of magnitude lower; more than 50 micrograms/ml of the protein was required for gelation. At 37 degrees C, gelation activity of actinogelin at low Ca2+ concentration was decreased to about a quarter of that at 20 degrees C, but this was still higher than that of gizzard alpha-actinin at 20 degrees C. Thus, role of actinogelin as an efficient and Ca2+-regulated cross-linker of microfilaments was substantiated.
mesh_terms: D000199:Actins / Q000378:metabolism*; D000818:Animals; D001704:Biopolymers; D002118:Calcium / Q000494:pharmacology; D005782:Gels; D008099:Liver / Q000378:metabolism; D008840:Microfilament Proteins / Q000378:metabolism* / Q000502:physiology; D008854:Microscopy, Electron; D011189:Potassium Chloride / Q000494:pharmacology; D011485:Protein Binding; D051381:Rats; D013050:Spectrometry, Fluorescence; D013696:Temperature; D014783:Viscosity
chemical_list: D000199:Actins; D001704:Biopolymers; D005782:Gels; D008840:Microfilament Proteins; C029354:actinogelin; D011189:Potassium Chloride; D002118:Calcium