Purification of a stimulator of microtubule assembly from bovine thyroid.
A protein from bovine thyroid which stimulated the assembly of microtubules from phosphocellulose-purified tubulin was isolated. The purification scheme involved fractionation of bovine thyroid homogenates successively on phosphocellulose, DEAE-cellulose, and Sephadex G-200. Activity was determined as ability to stimulate phosphocellulose-purified tubulin assembly measured turbidimetrically. Purification of stimulatory activity resulted in the selective enrichment to near homogeneity of a 40,000-45,000-dalton protein. The molecular weight was estimated by Sephadex G-200 gel filtration and polyacrylamide gel electrophoresis. This protein stimulated both the rate and the extent of assembly of thyroid and brain tubulin. In the turbidity assay which contained 4 M glycerol, the threshold for assembly of brain tubulin (at 1.7 mg/ml) was 0.01 mg/ml for the thyroid stimulator. Brain and thyroid microtubules assembled with thyroid stimulator appeared normal in diameter and shape by electron microscopy. Thyroid stimulator activity was destroyed by trypsinization or boiling. Activity was maximal at 1 mM GTP and 4 mM MgCl2 and was blocked by colchicine. No stimulatory protein of similar molecular weight could be isolated from bovine brain by similar techniques.
D000818:Animals; D001921:Brain / Q000378:metabolism*; D002417:Cattle; D007700:Kinetics; D008854:Microscopy, Electron; D008869:Microtubule-Associated Proteins; D008870:Microtubules / Q000187:drug effects / Q000648:ultrastructure*; D008970:Molecular Weight; D009419:Nerve Tissue Proteins / Q000302:isolation & purification / Q000378:metabolism; D011506:Proteins / Q000302:isolation & purification* / Q000378:metabolism; D013961:Thyroid Gland / Q000502:physiology*; D014404:Tubulin / Q000378:metabolism
D008869:Microtubule-Associated Proteins; D009419:Nerve Tissue Proteins; D011506:Proteins; D014404:Tubulin