Allele-specific activators and inhibitors for kinesin.
Members of the kinesin superfamily are force-generating ATPases that drive movement and influence cytoskeleton organization in cells. Often, more than one kinesin is implicated in a cellular process, and many kinesins are proposed to have overlapping functions. By using conventional kinesin as a model system, we have developed an approach to activate or inhibit a specific kinesin allele in the presence of other similar motor proteins. Modified ATP analogs are described that do not activate either conventional kinesin or another superfamily member, Eg5. However, a kinesin allele with Arg-14 in its nucleotide binding pocket mutated to alanine can use a subset of these nucleotide analogs to drive microtubule gliding. Cyclopentyl-ATP is one such analog. Cyclopentyl-adenylylimidodiphosphate, a nonhydrolyzable form of this analog, inhibits the mutant allele in microtubule-gliding assays, but not wild-type kinesin or Eg5. We anticipate that the incorporation of kinesin mutants and allele-specific activators and inhibitors in in vitro assays should clarify the role of individual motor proteins in complex cellular processes.
Kapoor
T M
TM
Department of Cell Biology, Harvard Medical School, Boston, MA 02115, USA. Tarun_Kapoor@hms.harvard.edu
Mitchison
T J
TJ
D000244:Adenosine Diphosphate / Q000378:metabolism; D000255:Adenosine Triphosphate / Q000031:analogs & derivatives / Q000378:metabolism / Q000494:pharmacology*; D000266:Adenylyl Imidodiphosphate / Q000031:analogs & derivatives / Q000378:metabolism / Q000494:pharmacology*; D001665:Binding Sites; D019281:Dimerization; D006801:Humans; D016547:Kinesins / Q000737:chemistry / Q000187:drug effects* / Q000378:metabolism*; D007700:Kinetics; D046911:Macromolecular Substances; D008958:Models, Molecular; D016297:Mutagenesis, Site-Directed; D011994:Recombinant Proteins / Q000737:chemistry / Q000187:drug effects / Q000378:metabolism; D013329:Structure-Activity Relationship
D046911:Macromolecular Substances; D011994:Recombinant Proteins; D000266:Adenylyl Imidodiphosphate; D000244:Adenosine Diphosphate; D000255:Adenosine Triphosphate; D016547:Kinesins