Configuration of flagellar microtubule subunits

Microtubule protofilaments and their subunits isolated from sperm flagellar doublet tubules of the sea urchin Strongylocentrotus purpuratus were examined by analytical biochemistry and high-resolution negative staining electron microscopy. All microtubule (tubulin) fractions show 2 polypeptide bands (α and β tubulins) in an approximate 1:1 ratio on urea-polyacryl-amide gel electrophoresis. Heat (37 °C)-solubilized microtubules yield a protein fraction containing the tubulin dimer of molecular weight 115 000 Daltons. The dimeric tubulin subunit, as seen in the electron microscope, has an overall size of about 3·5 × 8 nm and appears to have the configuration of a figure 8 because of stain penetration into the centre of each of its 2 halves (figure os). Isolated protofilaments (3·5 ± 0·3 nm thick) can each be resolved into 2 subfilaments (1·7 ± 0·2 nm thick). The 2 subfilaments have periodic lateral associations resulting in the basic 4-nm subunit repeat (figure o) along the protofilament. Examination of collapsed and solubilizing protofilaments shows the figure 8 (dimeric) subunits separating at random along the proto-filaments. We conclude that the tubulin dimer must be composed of either elongated or bilobed monomers which result in the figure 8 configuration and hence the 2-stranded appearance of the protofilaments.