Purification and characterization of kinesin from bovine adrenal medulla.
Kinesin was purified from bovine adrenal medulla. The sedimentation coefficient was 8.8 S. Sedimentation equilibrium ultracentrifugation studies showed the molecular weight of kinesin to be 300,000. The calculated axial ratio was 1:16. The Stokes radius was estimated to be 8.9 nm by gel filtration. Circular dichroism showed the alpha-helix content to be about 50%. Purified kinesin preparation contained a major polypeptide with a molecular weight of 120,000 and minor ones with molecular weights of 71,000, 68,000, and 65,000. Bovine adrenal kinesin had an ATPase activity which was stimulated severalfold by microtubules to a specific activity of about 0.1 mumol/min.mg. Kinesin molecules adsorbed to a glass slide promoted the movement of microtubules on the glass surface at a rate of about 0.5 micron/s. Immunostaining of EBTr (bovine embryonic trachea fibroblast) cells and bovine adrenal chromaffin cells in interphase with an affinity-purified antibody against the major polypeptide of kinesin showed that some kinesin was located on microtubules and the rest distributed throughout the cytoplasm in a diffuse manner. EBTr cells in mitotic phase gave a staining pattern showing that kinesin was present throughout the cytoplasm with higher concentration in the region of mitotic apparatus.
Murofushi
H
H
Department of Biophysics and Biochemistry, Faculty of Science, University of Tokyo, Japan.
Ikai
A
A
Okuhara
K
K
Kotani
S
S
Aizawa
H
H
Kumakura
K
K
Sakai
H
H
D000251:Adenosine Triphosphatases / Q000378:metabolism; D000313:Adrenal Medulla / Q000032:analysis*; D000327:Adsorption; D000818:Animals; D017301:Ca(2+) Mg(2+)-ATPase / Q000378:metabolism; D000252:Calcium-Transporting ATPases / Q000378:metabolism; D002417:Cattle; D002478:Cells, Cultured; D055598:Chemical Phenomena; D002627:Chemistry, Physical; D002850:Chromatography, Gel; D002942:Circular Dichroism; D005898:Glass; D007118:Immunoassay; D016547:Kinesins; D008868:Microtubule Proteins; D008870:Microtubules / Q000502:physiology; D008970:Molecular Weight; D009419:Nerve Tissue Proteins / Q000302:isolation & purification* / Q000378:metabolism; D011487:Protein Conformation; D014461:Ultracentrifugation
D008868:Microtubule Proteins; D009419:Nerve Tissue Proteins; D000251:Adenosine Triphosphatases; D017301:Ca(2+) Mg(2+)-ATPase; D016547:Kinesins; D000252:Calcium-Transporting ATPases