Gard D L, Kirschner M W
Department of Biochemistry and Biophysics, University of California School of Medicine, San Francisco 94143.
J Cell Biol. 1987 Nov;105(5):2203-15. doi: 10.1083/jcb.105.5.2203.
We have isolated a protein factor from Xenopus eggs that promotes microtubule assembly in vitro. Assembly promotion was associated with a 215-kD protein after a 1,000-3,000-fold enrichment of activity. The 215-kD protein, termed Xenopus microtubule assembly protein (XMAP), binds to microtubules with a stoichiometry of 0.06 mol/mol tubulin dimer. XMAP is immunologically distinct from the Xenopus homologues to mammalian brain microtubule-associated proteins; however, protein species immunologically related to XMAP with different molecular masses are found in Xenopus neuronal tissues and testis. XMAP is unusual in that it specifically promotes microtubule assembly at the plus-end. At a molar ratio of 0.01 mol XMAP/mol tubulin the assembly rate of the microtubule plus-end is accelerated 8-fold while the assembly rate of the minus-end is increased only 1.8-fold. Under these conditions XMAP promotes a 10-fold increase in the on-rate constant (from 1.4 s-1.microM-1 for microtubules assembled from pure tubulin to 15 s-1.microM-1), and a 10-fold decrease in off-rate constant (from 340 to 34 s-1). Given its stoichiometry in vivo, XMAP must be the major microtubule assembly factor in the Xenopus egg. XMAP is phosphorylated during M-phase of both meiotic and mitotic cycles, suggesting that its activity may be regulated during the cell cycle.
我们从非洲爪蟾卵中分离出一种蛋白质因子，它能在体外促进微管组装。在活性富集1000至3000倍后，组装促进作用与一种215-kD的蛋白质相关。这种215-kD的蛋白质被称为非洲爪蟾微管组装蛋白（XMAP），它与微管结合的化学计量比为0.06摩尔/摩尔微管蛋白二聚体。XMAP在免疫学上与哺乳动物脑微管相关蛋白的非洲爪蟾同源物不同；然而，在非洲爪蟾的神经组织和睾丸中发现了与XMAP免疫相关但分子量不同的蛋白质种类。XMAP的独特之处在于它能特异性地促进微管正端的组装。当XMAP与微管蛋白的摩尔比为0.01摩尔时，微管正端的组装速率加快8倍，而负端的组装速率仅增加1.8倍。在这些条件下，XMAP使结合速率常数增加10倍（从由纯微管蛋白组装的微管的1.4 s-1·μM-1增加到15 s-1·μM-1），解离速率常数降低10倍（从340降至34 s-1）。鉴于其在体内的化学计量比，XMAP必定是非洲爪蟾卵中的主要微管组装因子。在减数分裂和有丝分裂周期的M期，XMAP会发生磷酸化，这表明其活性可能在细胞周期中受到调控。
