Analysis of heterodimer formation by Xklp3A/B, a newly cloned kinesin-II from Xenopus laevis.
kinesin-II motor proteins are composed of two different kinesin-like motor proteins and one cargo binding subunit. Here we report the cloning of a new member of the kinesin-II superfamily, Xklp3A from Xenopus laevis, which forms a heterodimeric complex with Xklp3B. The heterodimer formation properties between Xklp3A and B have been tested in vitro using reticulocyte lysate expression and immunoprecipitation. To this end we produced a series of Xklp3A and B constructs of varying length and tested their propensity for heterodimer formation. We could demonstrate that, in contrast to conventional kinesin, the critical domains for heterodimer formation in Xklp3A/B are located at the C-terminal end of the stalk. Neither the neck nor the highly charged stretches after the neck region, which are typical of kinesins-II, are required for heterodimer formation, nor do they prevent homodimer formation. Dimerization is controlled by a cooperative mechanism between the C-terminal coiled-coil segments. Classical trigger sites were not identified. The critical regions for dimerization exhibit a very high degree of sequence conservation among equivalent members of the kinesin-II family.
De Marco
V
V
EMBL Structural and Computational Biology Programme, Meyerhofstrasse 1, 69117 Heidelberg, Germany.
Burkhard
P
P
Le Bot
N
N
Vernos
I
I
Hoenger
A
A
D000595:Amino Acid Sequence; D000818:Animals; D002135:Calcium-Binding Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D003001:Cloning, Molecular; D017124:Conserved Sequence; D019281:Dimerization; D016547:Kinesins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009124:Muscle Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D011487:Protein Conformation; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D020449:Repetitive Sequences, Amino Acid; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D029867:Xenopus Proteins; D014982:Xenopus laevis
D002135:Calcium-Binding Proteins; C499377:KIF3A protein, Xenopus; D009124:Muscle Proteins; D011994:Recombinant Proteins; C499376:XKLP3 protein, Xenopus; D029867:Xenopus Proteins; C074152:kinesin-II; D016547:Kinesins