Filament assembly from profilin-actin.
Profilin plays a major role in the assembly of actin filament at the barbed ends. The thermodynamic and kinetic parameters for barbed end assembly from profilin-actin have been measured turbidimetrically. Filament growth from profilin-actin requires MgATP to be bound to actin. No assembly is observed from profilin-CaATP-actin. The rate constant for association of profilin-actin to barbed ends is 30% lower than that of actin, and the critical concentration for F-actin assembly from profilin-actin units is 0.3 microM under physiological ionic conditions. Barbed ends grow from profilin-actin with an ADP-Pi cap. Profilin does not cap the barbed ends and is not detectably incorporated into filaments. The EDC-cross-linked profilin-actin complex (PAcov) both copolymerizes with F-actin and undergoes spontaneous self-assembly, following a nucleation-growth process characterized by a critical concentration of 0.2 microM under physiological conditions. The PAcov polymer is a helical filament that displays the same diffraction pattern as F-actin, with layer lines at 6 and 36 nm. The PAcov filaments bound phalloidin with the same kinetics as F-actin, bound myosin subfragment-1, and supported actin-activated ATPase of myosin subfragment-1, but they did not translocate in vitro along myosin-coated glass surfaces. These results are discussed in light of the current models of actin structure.
Laboratoire d'Enzymologie et Biochimie Structurales, CNRS, 91198 Gif-sur-Yvette, France.
I
I
Gutsche-Perelroizen
J
J
Lepault
A
A
Ott
M F
MF
Carlier
D008841:Actin Cytoskeleton / Q000737:chemistry* / Q000648:ultrastructure; D000199:Actins / Q000737:chemistry* / Q000648:ultrastructure; D000818:Animals; D003285:Contractile Proteins; D019281:Dimerization; D007700:Kinetics; D008840:Microfilament Proteins / Q000737:chemistry* / Q000648:ultrastructure; D051304:Profilins; D011485:Protein Binding; D011487:Protein Conformation; D011817:Rabbits; D013816:Thermodynamics
D000199:Actins; D003285:Contractile Proteins; D008840:Microfilament Proteins; D051304:Profilins