X-ray diffraction from microtubules.

X-ray diffraction patterns from sperm-tail microtubules indicate that subunits with a 40 to 50 A packing diameter form filaments, alternately half-staggered, parallel to the tubule axis. A 12-or a 13-stranded structure fits best with the X-ray diagram. The strongest bonding is that between units within a longitudinal filament; the weaker lateral interactions are disrupted by drying. Microtubules are the structural elements of many motile systems. In particular, cilia and flagella have a " 9 + 2 " arrangement of these tubules, which appear in the nine " outer fibers " as connected pairs of hollow cylinders, each about 240 A in diameter (Gibbons t Grimstone, 1960). Electron microscopic studies of the negatively stained, intact structure reveal subunits corresponding to the globular protein of molecular weight 60,000 obtained from disaggregated microtubules Analysis of optical diffraction patterns of electron micrographs from Trichonympha flagella indicates a helical arrangement of subunits in the surface lattice of the microtubules, with a cell 40 b, x 50 d (Grimstone & Klug, 1966). In this paper we describe an X-ray diffraction study of hydrated microtubules. Previous X-ray work on unpurified cilia and sperm tails reported diffractions from the membrane lipid phase only (Pautard, 1952; Silvester, 1964). Our preparations of gels of sperm-tail outer fibers are weakly diffracting, and the diagram is not detailed. Nevertheless, the patterns reveal a surface lattice somewhat different from the one seen in the electron microscope, and we can place limits on possible symmetries for the structure. The simplest model, is a 12-or a 13-stranded array, with neighboring strands approximately half-staggered. Sperm tails from Strongylocentrotus droebachiensis were prepared by the method of Stephens, Renaud & Gibbons (1967). Three different procedures were used to remove the membranes: treatment with digitonin (Gibbons, 1965), washing with 1% Triton X-1000 (Stephens & Linck, 1969), and extraction with 50% glycerol for two days at 0°C. (The latter procedures were introduced after we observed that removal of membranes with digitonin produced a digitonin-cholesterol complex that contaminated subsequent " purified " samples. Strong diffraction from this complex was observed at 5.1 d and 6.1 A and in the 50 A region. These spacings have been erroneously attributed (Forslind, Swanbeck & Mohri, 1968) to diffraction from the microtubules themselves.) Purification of outer fibers then proceeded by dialysis (Stephens et al., 1967). For some experiments, A-subfibers were prepared by thermal depolymerization of the B-subfibers at low ionic strength (Stephens, 1970). Suspensions of …