Sale W S, Gibbons I R
J Cell Biol. 1979 Jul;82(1):291-8. doi: 10.1083/jcb.82.1.291.
The effect of vanadate on the ATP-induced disruption of trypsin-treated axonemes and the ATP-induced straightening of rigor wave preparations of sea urchin sperm was investigated. Addition of ATP to a suspension of trypsin-treated axonemes results in a rapid decrease in turbidity (optical density measured at 350 nm) concomitant with the disruption of the axonemes by sliding between microtubules to form tangles of connected doublet microtubules (Summers and Gibbons, 1971; Sale and Satir, 1977). For axonemes digested to approximately 93 percent of their initial turbidity, 5 {muM} vanadate completely inhibits the ATP-induced decrease in turbidity and the axonemes maintain their structural integrity. However, with axonemes digested to approximately 80 percent of their initial turbidity, vanadate fails to inhibit the ATP-induced decrease in turbidity and the ATP-induced structural disruption of axonemes, even when the vanadate concentration is raised as high as 100 mum. For such axonemes digested to 80 percent of their initial turbidity, the form of ATP-induced structural changes, in the presence of 25 muM vanadate, was observed by dark-field light microscopy and revealed that the axonemes become disrupted into curved, isolated doublet microtubules, small groups of doublet microtubules, and "banana peel" structures in which tubules have peeled back from the axoneme. Addition of 5 muM ATP to rigor wave sperm, which were prepared by abrupt removal of ATP from reactivated sperm, causes straightening of the rigor waves within 1 min, and addition of more than 10 muM ATP causes resumption of flagellar beating. Addition of 40 muM vanadate to the rigor wave sperm does not inhibit straightening of the rigor waves of 2 muM-1 mM ATP, although oscillatory beating is completely inhibited. These results suggest that vanadate inhibits the mechanochemical cycle of dyein at a step subsequent to the MgATP(2-)-induced release of the bridged dynein arms.
研究了钒酸盐对ATP诱导的经胰蛋白酶处理的轴丝破坏以及ATP诱导的海胆精子强直波制剂伸直的影响。向经胰蛋白酶处理的轴丝悬浮液中添加ATP会导致浊度迅速降低（在350nm处测量光密度），同时轴丝通过微管间滑动而破坏，形成相连双联体微管的缠结（萨默斯和吉本斯，1971；塞尔和萨蒂尔，1977）。对于消化至初始浊度约93%的轴丝，5μM钒酸盐完全抑制ATP诱导的浊度降低，轴丝保持其结构完整性。然而，对于消化至初始浊度约80%的轴丝，即使钒酸盐浓度提高至100μM，钒酸盐也无法抑制ATP诱导的浊度降低和轴丝的ATP诱导的结构破坏。对于消化至初始浊度80%的此类轴丝，在25μM钒酸盐存在下，通过暗视野光学显微镜观察ATP诱导的结构变化形式，发现轴丝被破坏成弯曲的、孤立的双联体微管、小群双联体微管以及“香蕉皮”结构，其中微管从轴丝上剥离。向通过从再激活的精子中突然去除ATP制备的强直波精子中添加5μM ATP会在1分钟内导致强直波伸直，添加超过10μM ATP会导致鞭毛重新开始摆动。向强直波精子中添加40μM钒酸盐不会抑制2μM - 1mM ATP诱导的强直波伸直，尽管振荡摆动被完全抑制。这些结果表明，钒酸盐在MgATP(2 - )诱导桥接动力蛋白臂释放之后的步骤抑制动力蛋白的机械化学循环。
