Interpreting a medium-resolution model of tubulin: comparison of zinc-sheet and microtubule structure.
We previously used electron crystallography of zinc-induced two-dimensional crystalline sheets of tubulin to construct a medium-resolution three dimensional (3-D) reconstruction (at 6.5 A) of this protein. Here we present an improved model, and extend the interpretation to correlate it to microtubule structure. Secondary sequence predictions and projection density maps of subtilisin-cleaved tubulin provide information on the location of the C-terminal portion, which has been suggested to be involved in the binding of microtubule-associated proteins. The zinc-sheet tubulin model is compared to microtubules in two ways; comparison of electron diffraction from the zinc-sheets to electron diffraction from microtubules, and by docking the zinc-sheet protofilament 3-D model into a helical reconstruction from ice-embedded microtubules. By correlating the zinc-sheet protofilament to a reconstruction of axonemal protofilaments, we assigned polarity to the protofilament in our model. The polarity assignment together with our model for dimer boundaries and the assignment of alpha- and beta-monomers in our reconstruction, provides a microtubule model where the alpha-monomer crowns the plus- (or fast-growing) end of the microtubule and contact is made in the centrosome with gamma-tubulin via the beta-monomer.
Life Sciences Division, Lawrence Berkeley National Laboratory, CA 94720, USA.
S G
SG
Wolf
E
E
Nogales
M
M
Kikkawa
D
D
Gratzinger
N
N
Hirokawa
K H
KH
Downing
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D002417:Cattle; D003460:Crystallization; D006160:Guanosine Triphosphate / Q000378:metabolism; D008854:Microscopy, Electron; D008870:Microtubules / Q000737:chemistry* / Q000648:ultrastructure; D008958:Models, Molecular; D008969:Molecular Sequence Data; D017239:Paclitaxel / Q000378:metabolism; D011487:Protein Conformation; D017433:Protein Structure, Secondary; D013381:Subtilisins / Q000737:chemistry; D014404:Tubulin / Q000737:chemistry* / Q000648:ultrastructure; D015032:Zinc
D014404:Tubulin; D006160:Guanosine Triphosphate; D013381:Subtilisins; D015032:Zinc; D017239:Paclitaxel