Low resolution structure of microtubules in solution. Synchrotron X-ray scattering and electron microscopy of taxol-induced microtubules assembled from purified tubulin in comparison with glycerol and MAP-induced microtubules.
The structure of microtubules has been characterized to 3 nm resolution employing time-resolved X-ray scattering. This has revealed detailed structural features of microtubules not observed before in solution. The polymerization of highly purified tubulin, induced by the antitumour drug taxol, has been employed as a microtubule model system. This assembly reaction requires Mg2+, is optimal at a 1:1 taxol to tubulin heterodimer molar ratio, proceeds with GTP or GDP and is intrinsically reversible. The X-ray scattering profiles are consistent with identical non-globular alpha and beta-tubulin monomers ordered within the known helical surface lattice of microtubules. Purified tubulin-taxol microtubules have a smaller mean diameter (approx. 22 nm) than those induced by microtubule associated proteins or glycerol (approx. 24 nm), but nearly identical wall substructure to the resolution of the measurements. This is because the majority of the former consist of only 12 protofilaments instead of the typical 13 protofilaments, as confirmed by electron microscopy of thin-sectioned, negatively stained and ice-embedded taxol microtubules. It may be concluded that taxol induces a slight reduction of the lateral contact curvature between tubulin monomers. The main fringe pattern observed in cryo-electron micrographs is consistent with a simple 12 protofilament 3-start skewed lattice model. Cylindrical closure of this lattice can be achieved by tilting the lattice 0.8 degrees with respect to the microtubule axis. The closure implies a discontinuity in the type of lateral contacts between the tubulin monomers (regardless of whether these are of the -alpha-beta- or the -alpha-alpha-/-beta-beta- type), which indicates that lateral contacts and the subunit specificity of taxol binding are, to a large degree, equivalent.
Centro de Investigaciones Biológicas, C.S.I.C., Madrid, Spain.
J M
JM
Andreu
J
J
Bordas
J F
JF
Diaz
J
J
García de Ancos
R
R
Gil
F J
FJ
Medrano
E
E
Nogales
E
E
Pantos
E
E
Towns-Andrews
D000470:Alkaloids / Q000494:pharmacology*; D005990:Glycerol; D006150:Guanine Nucleotides / Q000378:metabolism; D008274:Magnesium / Q000378:metabolism; D008854:Microscopy, Electron; D008869:Microtubule-Associated Proteins / Q000378:metabolism; D008870:Microtubules / Q000737:chemistry / Q000187:drug effects* / Q000648:ultrastructure; D017239:Paclitaxel; D012542:Scattering, Radiation; D014404:Tubulin / Q000378:metabolism*; D014965:X-Rays
D000470:Alkaloids; D006150:Guanine Nucleotides; D008869:Microtubule-Associated Proteins; D014404:Tubulin; D008274:Magnesium; D017239:Paclitaxel; D005990:Glycerol