A large body of biochemical evidence suggests that the F-actin filament can have internal cooperativity. We have observed large cooperative effects on the low-resolution structure of actin filaments under three very different conditions. First, when G-Ca2+-actin is polymerized by both Mg2+and KCl, filaments may be found in two different populations, with two discrete positions seen for subdomain 2. When G- Ca2+actin is polymerized by only Mg2+, a single F-Mg2+-actin population is seen. The structural data suggest that an entire filament exists with subdomain 2 in one state or the other when there is a heterogenous mixture of Mg2+and Ca2+-actin. Second, when actin filaments are nucleated from gelsolin there is a conformational change that can be observed throughout the filament that is consistent with a large shift in the actin C terminus. There must be a large cooperative propagation of this effect throughout the filament from the nucleation piont. Third, we have used phalloidin to stabilize F-actin in which two C-terminal residues have been proteolytically removed by trypsin. It has been shown biochemically that this stabilization occurs at substoichometric amounts of phalloidin. Phalloidin, at either a 1:1 or a 1:20 molar ratio with actin, restores the connectivity between the long-pitch helical strands. F-actin's internal cooperativity will have large implicationsin vivo, particularly in muscle.
Bibliographical noteFunding Information:
We thank Dr Paul Janmey for the very generous gift of gelsolin, and Brooks Rogers for assistance with the image analysis of gelsolin-nucleated filaments. We thank Dr David Thomas for helpful discussions. This work was supported by NIH AR42023 (to E.H.E.), AR32961 (to David Thomas) and a grant from the American Heart Association Minnesota Affiliate (to E.P.).
- Electron microscopy
- Image analysis
- Protein polymers