1: Biophys J  1996 Jan;70(1):401-8 

Preparation of bead-tailed actin filaments: estimation of the torque produced by
the sliding force in an in vitro motility assay.

Suzuki N, Miyata H, Ishiwata S, Kinosita K Jr.

Department of Physics, Faculty of Science and Technology, Keio University,
Yokohama, Japan.

By coating covalently the surface of a polystyrene bead (diameter = 1 micron)
with gelsolin, we have succeeded in attaching the bead selectively at the barbed
end of an actin filament and forming a 1:1 bead-actin filament complex. On a
layer of heavy meromyosin on a nitrocellulose-coated coverglass, this bead-actin
filament complex slid smoothly, trailing the bead at its end. Therefore we
called this preparation "bead-tailed" actin filaments. The sliding velocity was
indistinguishable from that of nonbeaded filaments. With use of this system, we
tried to detect the axial rotation (rotation around the filament axis) in a
sliding actin filament. Although a single bead at the tail end did not serve as
the marker for the axial rotation, we occasionally found another bead bound to
the tail bead. In this case, the orientation of the bead-aggregate could be
followed continuously with a video monitor while the filament was sliding over
heavy meromyosin. We observed that actin filaments slid over distances of many
tens of micrometers without showing a complete turn of the bead-aggregates. On
the basis of the calculation of rotational friction drag on the bead-aggregate,
we estimate that the rotational component of the sliding force and the torque
produced on a sliding actin filament (length < or = 10 microns) did not
accumulate > 1 pN and 5 pN.nm, respectively. In the present system of randomly
oriented heavy meromyosin lying on a nitrocellulose film without an external
load.

PMID: 8770216 [PubMed - indexed for MEDLINE]