Elongation of actin filaments is a diffusion-limited reaction at the barbed end and is accelerated by inert macromolecules.
We used a fluorescence method to measure the rate constants for the elongation of pyrene-labeled actin filaments in a number of different solvents. The absolute values of the rate constants were established by electron microscopy. Using glycerol, sucrose, or ethylene glycol to vary the solution viscosity, the association rate constant (k+) was 10(7) M-1 s-1 viscosity-1 (in centipoise). Consequently, plots of 1/k+ versus viscosity are linear and extrapolate to near the origin as expected for a diffusion-limited reaction where the rate constant approaches infinity at zero viscosity. By electron microscopy, we found that this inhibitory effect of glycerol is almost entirely at the fast growing, barbed end. For the pointed end, plots of 1/k+ versus viscosity extrapolate to a maximum rate of about 10(6) M-1 s-1 at zero viscosity, so that elongation at the pointed is not limited by diffusion. In contrast to these small molecules, polyethylene glycol, dextran, and ovalbumin all cause a concentration (and therefore viscosity)-dependent increase in k+. At any given viscosity, their effects are similar to each other. For example, at 3 centipoise, k+ = 2.2 X 10(7) M-1 s-1. We presume that this is due to an excluded volume effect that causes an increase in the thermodynamic activity of the actin. If the proteins in the cytoplasmic matrix have a similar effect, the association reactions of actin in cells may be much faster than expected from experiments done in dilute buffers.
D000199:Actins / Q000378:metabolism*; D000818:Animals; D004058:Diffusion; D019855:Ethylene Glycol; D005026:Ethylene Glycols / Q000494:pharmacology; D005990:Glycerol / Q000494:pharmacology; D008433:Mathematics; D008854:Microscopy, Electron; D011108:Polymers / Q000378:metabolism; D011817:Rabbits; D013050:Spectrometry, Fluorescence; D013395:Sucrose / Q000494:pharmacology; D013696:Temperature; D014783:Viscosity
D000199:Actins; D005026:Ethylene Glycols; D011108:Polymers; D013395:Sucrose; D019855:Ethylene Glycol; D005990:Glycerol