Kinesin and tau bind to distinct sites on microtubules.
We have used a fluorescent derivative of kinesin, AF-kinesin (kinesin conjugated with 5-(iodoacetamido)fluorescein), to investigate the binding site of kinesin on microtubules and to compare this site with that to which tau binds. Microtubules saturated with tau will bind AF-kinesin in the presence of the ATP analogue, 5'-[beta,gamma-imino]triphosphate (AdoPP[NH]P). This shows that there are distinct binding sites for the two proteins. Further evidence comes from digestion studies where taxol-stabilised microtubules were treated with subtilisin, resulting in the cleavage of C-terminal residues from both the alpha- and beta-tubulin subunits. These treated microtubules can no longer bind tau, but are able to bind AF-kinesin in the presence of AdoPP[NH]P. Finally, AF-kinesin will support the gliding of subtilisin-digested microtubules in the presence of ATP at rates comparable to those obtained with non-digested microtubules. These results show directly that the binding site for kinesin is outside the C-terminal region of tubulin that is removed by subtilisin and is distinct from the binding site of tau.
Marya
P K
PK
Department of Molecular Biology and Biophysics, Randall Institute, King's College, University of London, UK.
Syed
Z
Z
Fraylich
P E
PE
Eagles
P A
PA
D000266:Adenylyl Imidodiphosphate / Q000494:pharmacology; D000818:Animals; D001665:Binding Sites; D001921:Brain / Q000378:metabolism; D002417:Cattle; D002848:Chromatography, DEAE-Cellulose; D002851:Chromatography, High Pressure Liquid; D004591:Electrophoresis, Polyacrylamide Gel; D005452:Fluoresceins; D016547:Kinesins / Q000302:isolation & purification / Q000378:metabolism*; D008870:Microtubules / Q000378:metabolism* / Q000648:ultrastructure; D008970:Molecular Weight; D017239:Paclitaxel / Q000494:pharmacology; D014404:Tubulin / Q000302:isolation & purification / Q000378:metabolism*; D016875:tau Proteins / Q000302:isolation & purification / Q000378:metabolism*
D005452:Fluoresceins; D014404:Tubulin; D016875:tau Proteins; D000266:Adenylyl Imidodiphosphate; C015296:5-iodoacetamidofluorescein; D016547:Kinesins; D017239:Paclitaxel