Jameson L, Caplow M
Proc Natl Acad Sci U S A. 1981 Jun;78(6):3413-7. doi: 10.1073/pnas.78.6.3413.
Phosphorylation of purified microtubule-associated proteins (MAPs) inhibits the rate and extent of MAP-stimulated microtubule assembly. The extent of microtubule assembly is reduced as a result of a decrease in the fraction of tubulin polymerized, without a significant change in the critical protein concentration. The decreased rate of microtubule assembly using phosphorylated MAPs reflects a reduction in microtubule nucleation resulting in fewer, but 2-fold longer, microtubules at steady state. Analysis of microtubule (MT) dynamics at steady state reveals that the rate of directional incorporation of subunits (flux) is 22 subunits.MT-1.sec-1 with phosphorylated MAPs, compared to 10 subunits.MT-1.sec-1 with unphosphorylated MAPs. The initial rate of disassembly determined by isothermal dilution is 232 subunits.MT-1.sec-1 for microtubules assembled with phosphorylated MAPs, compared to 102 subunits.MT-1.sec-1 for microtubules assembled with unphosphorylated MAPs. By using these results, the directionality (the number of successful subunit additions relative to the total number of association events per unit time) for subunit addition is found to be 0.1 for microtubules assembled with either phosphorylated or unphosphorylated MAPs. These observations are interpreted in terms of a mechanism in which phosphorylation of MAPs increases the rate of steady-state subunit flux by an equivalent enhancement of the rates of subunit association and dissociation, such that the critical protein concentration and directionality remain unchanged.
纯化的微管相关蛋白（MAPs）的磷酸化会抑制MAPs刺激的微管组装速率和程度。微管组装程度降低是由于聚合的微管蛋白比例下降，而临界蛋白浓度没有显著变化。使用磷酸化MAPs时微管组装速率降低反映了微管成核减少，导致稳态下微管数量减少，但长度增加了2倍。对稳态下微管（MT）动力学的分析表明，亚基定向掺入（通量）的速率在磷酸化MAPs存在时为22个亚基·MT⁻¹·秒⁻¹，而在未磷酸化MAPs存在时为10个亚基·MT⁻¹·秒⁻¹。通过等温稀释测定的微管解聚初始速率，对于用磷酸化MAPs组装的微管为232个亚基·MT⁻¹·秒⁻¹，而对于用未磷酸化MAPs组装的微管为102个亚基·MT⁻¹·秒⁻¹。利用这些结果发现，对于用磷酸化或未磷酸化MAPs组装的微管，亚基添加的方向性（相对于每单位时间总结合事件数成功添加的亚基数量）均为0.1。这些观察结果可通过一种机制来解释，即MAPs的磷酸化通过同等程度地提高亚基结合和解离速率来增加稳态亚基通量速率，从而使临界蛋白浓度和方向性保持不变。
