Actin Monomers Assemble into Long Helical FActin Polymers

Actin exists as a globular monomer called G-actin and as a filamentous polymer called F-actin, which is a linear chain of G-actin subunits. (The microfilaments visualized in a cell by electron microscopy are F-actin filaments plus any bound proteins.) Each actin molecule contains a Mg2+ ion complexed with either ATP or ADP. Thus there are four states of actin: ATP-G-actin, ADP-G-actin, ATP-F-actin, and ADP-F-actin. Two of these forms, ATP-G-actin and ADP-F-actin, predominate in a cell. The importance of the interconversion between the ATP and the ADP forms of actin in the assembly of the cytoskeleton is discussed later.

Although G-actin appears globular in the electron microscope, x-ray crystallographic analysis reveals that it is separated into two lobes by a deep cleft (Figure 19-3a). The lobes and the cleft compose the ATPase fold, the site where ATP

(+) end

(9 nm diameter) (arrows). (c) In one model of the arrangement of subunits in an actin filament, the subunits lie in a tight helix along the filament, as indicated by the arrow. One repeating unit consists of 28 subunits (13 turns of the helix), covering a distance of 72 nm. Only 14 subunits are shown in the figure. The ATP-binding cleft is oriented in the same direction (top) in all actin subunits in the filament. The end of a filament with an exposed binding cleft is designated the (—)end; the opposite end is the ( + ) end. [Part (a) adapted from C. E. Schutt et al., 1993, Nature 365:810; courtesy of M. Rozycki. Part (b) courtesy of R. Craig. Part (c) see M. F. Schmid et al., 1994, J. Cell Biol. 124:341; courtesy of M. Schmid.]




C-terminus and Mg2+ are bound. In actin, the floor of the cleft acts as a hinge that allows the lobes to flex relative to each other. When ATP or ADP is bound to G-actin, the nucleotide affects the conformation of the molecule. In fact, without a bound nucleotide, G-actin denatures very quickly.

The addition of ions—Mg2 + , K+, or Na+—to a solution of G-actin will induce the polymerization of G-actin into F-actin filaments. The process is also reversible: F-actin depolymerizes into G-actin when the ionic strength of the solution is lowered. The F-actin filaments that form in vitro are indistinguishable from microfilaments isolated from cells, indicating that other factors such as accessory proteins are not required for polymerization in vivo. The assembly of G-actin into F-actin is accompanied by the hydrolysis of ATP to ADP and P(; however, as discussed later, ATP hydrolysis affects the kinetics of polymerization but is not necessary for polymerization to take place.

When negatively stained by uranyl acetate for electron microscopy, F-actin appears as twisted strings of beads whose diameter varies between 7 and 9 nm (Figure 19-3b). From the results of x-ray diffraction studies of actin filaments and the actin monomer structure shown in Figure 19-3a, scientists have produced a model of an actin filament in which the subunits are organized as a tightly wound helix (Figure 19-3c). In this arrangement, each subunit is surrounded by four other subunits, one above, one below, and two to one side. Each subunit corresponds to a bead seen in electron micrographs of actin filaments.

The ability of G-actin to polymerize into F-actin and of F-actin to depolymerize into G-actin is an important property of actin. In this chapter, we will see how the reversible assembly of actin lies at the core of many cell movements.

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