Domains of tau protein and interactions with microtubules.
The role of the neuronal microtubule-associated protein tau has been studied by generating a series of tau constructs differing in one or several of its subdomains: length and composition of the repeat domains, extensions of the repeats in the N- or C-terminal direction, constructs without repeats, assembly vs projection domain, and number of N-terminal inserts. The interaction of the mutant tau proteins with microtubules was judged by several independent methods. (i) Direct binding assays between tau and taxol-stabilized microtubules yield dissociation constants and stoichiometries. (ii) Light scattering and X-ray scattering of assembling microtubule solutions reflect the capacity of tau to promote microtubule nucleation, elongation, and bundling in bulk solution. (iii) Dark field microscopy of assembling microtubules allows one to assess the efficiency of nucleation and bundling separately. The repeat region alone, the N-terminal domains alone, or the C-terminal tail alone binds only weakly to microtubules. However, binding is strongly enhanced by combinations such as the repeat region plus one or both of the flanking regions which could be viewed as "jaws" for tau on the microtubule surface (the proline-rich domain P upstream of the repeats and the "fifth" repeat R' downstream). Such combinations make tau's binding productive in terms of microtubule assembly and stabilization, while the combination of the flanking regions without repeats binds only unproductively. Efficient nucleation parallels strong binding in most cases, i.e., when a construct binds tightly to microtubules, it also nucleates them efficiently and vice versa. In addition, the proline-rich domain P in combination with the repeats R or the flanking domain R' causes pronounced bundling. This effect disappears when the N-terminal domains (acidic or basic) are added on, suggesting that the tau isoforms are not "bundling proteins" in the proper sense. In spite of the wide range of binding strength and nucleation efficiency, the stoichiometries of binding are rather reproducible (around 0.5 tau/tubulin dimer); this is in remarkable contrast to the effect of certain types of phosphorylation which can strongly reduce the stoichiometry.
Gustke
N
N
Max-Planck-Unit for Structural Molecular Biology, Hamburg, Germany.
Trinczek
B
B
Biernat
J
J
Mandelkow
E M
EM
Mandelkow
E
E
D000595:Amino Acid Sequence; D004252:DNA Mutational Analysis; D006801:Humans; D008853:Microscopy / Q000379:methods; D008870:Microtubules / Q000378:metabolism* / Q000648:ultrastructure; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000378:metabolism*; D011485:Protein Binding; D011487:Protein Conformation; D011994:Recombinant Proteins / Q000378:metabolism; D012542:Scattering, Radiation; D013329:Structure-Activity Relationship; D014741:Video Recording; D014965:X-Rays; D016875:tau Proteins / Q000378:metabolism*
D010446:Peptide Fragments; D011994:Recombinant Proteins; D016875:tau Proteins