title: Flexibility of actin filaments derived from thermal fluctuations. Effect of bound nucleotide, phalloidin, and muscle regulatory proteins.
abstract: Single actin filaments undergoing brownian movement in two dimensions were observed at 20 degrees C in fluorescence optical video microscopy. The persistence length (Lp) was derived from the analysis of either the cosine correlation function or the average transverse fluctuations of a series of recorded shapes of filaments assembled from rhodamine-action. Phalloidin-stabilized filaments had a persistence length of 18 +/- 1 micron, in agreement with recent observations. In the absence of phalloidin, rhodamine-labeled filaments could be observed under a variety of solution conditions once diluted in free unlabeled G-actin at the appropriate critical concentration. Such nonstabilized F-ADP-actin filaments had the same Lp of 9 +/- 0.5 microns, whether they had been assembled from ATP-G-actin or from ADP-G-actin, and independently of the tightly bound divalent metal ion. In the presence of BeF3-, which mimics the gamma-phosphate of ATP, F-ADP-BeF3-actin was appreciably more rigid, with Lp = 13.5 microns. Hence, newly formed F-ADP-Pi-actin filaments are more rigid than "old" F-ADP-actin filaments, a fact which has implications in actin-based motility processes. In the presence of skeletal tropomyosin and troponin, filaments were rigid (Lp = 20 +/- 1 micron) in the off state (-Ca2+), and flexible (Lp = 12 microns) in the on state (+Ca2+), consistent with the steric blocking model. In agreement with x-ray diffraction data, no appreciable difference was recorded between the off and on states using smooth muscle tropomyosin and caldesmon (Lp = 20 +/- 1 micron). In conclusion, this method allows accurate measurement of small (< or = 15%) changes in mechanical properties of actin filaments in correlation with their biological functions.
authors.affiliation: Groupe de Physicochimie Théorique, Ecole Supérieure de Physique et Chimie Industrielles, Paris, France.
authors.forename: H
authors.initials: H
authors.lastname: Isambert
authors.forename: P
authors.initials: P
authors.lastname: Venier
authors.forename: A C
authors.initials: AC
authors.lastname: Maggs
authors.forename: A
authors.initials: A
authors.lastname: Fattoum
authors.forename: R
authors.initials: R
authors.lastname: Kassab
authors.forename: D
authors.initials: D
authors.lastname: Pantaloni
authors.forename: M F
authors.initials: MF
authors.lastname: Carlier
mesh_terms: D008841:Actin Cytoskeleton / Q000648:ultrastructure; D000199:Actins / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism*; D000244:Adenosine Diphosphate / Q000378:metabolism* / Q000494:pharmacology; D000255:Adenosine Triphosphate / Q000378:metabolism* / Q000494:pharmacology; D000818:Animals; D002148:Calmodulin-Binding Proteins / Q000378:metabolism; D006358:Hot Temperature; D008856:Microscopy, Fluorescence; D018482:Muscle, Skeletal / Q000378:metabolism; D010590:Phalloidine / Q000494:pharmacology; D011487:Protein Conformation; D011817:Rabbits; D013816:Thermodynamics; D014335:Tropomyosin / Q000378:metabolism*; D014336:Troponin / Q000378:metabolism*; D014961:X-Ray Diffraction
chemical_list: D000199:Actins; D002148:Calmodulin-Binding Proteins; D014335:Tropomyosin; D014336:Troponin; D010590:Phalloidine; D000244:Adenosine Diphosphate; D000255:Adenosine Triphosphate