© 2006 The Japanese Biochemical Society.
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Intermolecular Cross-Linking of a Novel Rice Kinesin K16 Motor Domain with a Photoreactive ATP Derivative
1 Laboratories of Plant and Microbial Genome Control, Graduate School of Science and Technology, Niigata University, Niigata 950-2181; and 2 Division of Bioengineering, Graduate School of Engineering, Soka University, Hachioji, Tokyo 192-8577
* To whom correspondence should be addressed: Fax: +81-426-91-9312, E-mail: shinsaku{at}t.soka.ac.jp (Shinsaku Maruta); Fax: +81-25-262-6641, E-mail: t.mitsui{at}agrews.agr.niigata-u.ac.jp(Toshiaki Mitsui)
A fluorescent photoreactive ATP derivative, 2'(3')-O-(4-benzoylbenzoyl)-1,N6-etheno-ATP (Bz2-
ATP), was synthesized and reacted with the rice kinesin K16 motor domain (K16MD). In the presence of ADP or ATP, UV irradiation of the K16MD solution containing Bz2-ATP resulted in a new 100 kDa band, which was an intermolecular cross-linked product of motor domains. In contrast, no cross-linking was observed in the absence of nucleotides. For the motor domain of mouse brain kinesin and skeletal muscle myosin subfragment-1, no such intermolecular photo cross-linking by Bz2-ATP was observed. Our results indicate that Bz2-ATP acts unusually as a photoreactive crosslinker to detect conformational changes in K16MD induced by nucleotide binding resulting in the formation of dimers.