Repeat motifs of tau bind to the insides of microtubules in the absence of taxol.
The tau family of microtubule-associated proteins has a microtubule-binding domain which includes three or four conserved sequence repeats. Pelleting assays show that when tubulin and tau are co- assembled into microtubules, the presence of taxol reduces the amount of tau incorporated. In the absence of taxol, strong binding sites for tau are filled by one repeat motif per tubulin dimer; additional tau molecules bind more weakly. We have labelled a repeat motif with nanogold and used three-dimensional electron cryomicroscopy to compare images of microtubules assembled with labelled or unlabelled tau. With kinesin motor domains bound to the microtubule outer surface to distinguish between alpha- and beta-tubulin, we show that the gold label lies on the inner surface close to the taxol binding site on beta-tubulin. Loops within the repeat motifs of tau have sequence similarity to an extended loop which occupies a site in alpha-tubulin equivalent to the taxol-binding pocket in beta-tubulin. We propose that loops in bound tau stabilize microtubules in a similar way to taxol, although with lower affinity so that assembly is reversible.
Kar
Santwana
S
MRC Laboratory of Molecular Biology, Hills Road, Cambridge CB2 2QH, UK.
Fan
Juan
J
Smith
Michael J
MJ
Goedert
Michel
M
Amos
Linda A
LA
D000595:Amino Acid Sequence; D003001:Cloning, Molecular; D006801:Humans; D007091:Image Processing, Computer-Assisted; D007700:Kinetics; D008870:Microtubules / Q000187:drug effects / Q000378:metabolism* / Q000648:ultrastructure; D008958:Models, Molecular; D008969:Molecular Sequence Data; D017239:Paclitaxel / Q000494:pharmacology*; D011487:Protein Conformation; D011994:Recombinant Proteins / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism; D020449:Repetitive Sequences, Amino Acid; D014404:Tubulin / Q000378:metabolism*; D016875:tau Proteins / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism*
D011994:Recombinant Proteins; D014404:Tubulin; D016875:tau Proteins; D017239:Paclitaxel