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.