KMID : 0380220070400010044
|
|
Journal of Biochemistry and Molecular Biology 2007 Volume.40 No. 1 p.44 ~ p.52
|
|
Two Kinesins from Arabidopsis, KatB and KatC, Have a Second Microtubule-binding Site in the Tail Domain
|
|
Jiang Shiling
Li Ming Xu Tao Ren Dongtao Liu Guoqin
|
|
Abstract
|
|
|
Kinesins, as a kind of microtubule-based motor proteins, have a conserved microtubule-binding site in their motor domain. Here we report that two homologous kinesins in Arabidopsis thaliana, KatB and KatC, contain a second microtubule-binding site in their tail domains. The prokaryotic-expressed N-terminal tail domain of the KatC heavy chain can bind to microtubules in an ATP-insensitive manner. To identify the precise region responsible for the binding, a serious of truncated KatC cDNAs encoding KatC N-terminal regions in different lengths, KatC1-128, KatC1-86, KatC1-73 and KatC1-63, fused to Histidine-tags, were expressed in E. coli and affinity-purified. Microtubule cosedimentation assays show that the site at amino acid residues 74-86 in KatC is important for microtubulebinding. By similarity, we obtained three different lengths of KatB N-terminal regions, KatB1-384, KatB1-77, and KatB1-63, and analyzed their microtubule-binding ability. Cosedimentation assays indicate that the KatB tail domain can also bind to microtubules at the same site as and in a similar manner to KatC. Fluorescence microscopic observations show that the microtubule-binding site at the tail domain of KatB or KatC can induce microtubules bundling only when the stalk domain is present. Through pull-down assays, we show that KatB1-385 and KatC1-394 are able to interact specifically with themselves and with each other in vitro. These findings are significant for identifying a previously uncharacterized microtubule-binding site in the two kinesin proteins, KatB and KatC, and the functional relations between them.
|
|
KEYWORD
|
|
Cosedimentation, Kinesin, KatB, KatC, Microtubule-binding site
|
|
FullTexts / Linksout information
|
|
|
|
Listed journal information
|
|
|
|