Monoclonal Antibodies to the [alpha]- and [beta]-Subunits of the Plant Mitochondrial F1-ATPase
Monoclonal Antibodies to the [alpha]- and [beta]-Subunits of the Plant Mitochondrial F1-ATPase
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植物线粒体 F1-ATP 酶 α- 和 β- 亚基的单克隆抗体
DOI：
10.1104/pp.101.3.931
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发表时间：
1993
期刊：
影响因子：
--
通讯作者：
T. Elthon
中科院分区：
文献类型：
--
作者：
M. Luethy;A. Horak;T. Elthon
我们已经制备了9株抗玉米（Zea mays L.）线粒体F1-ATPase。这些单克隆抗体通过针对玉米线粒体组分免疫小鼠并随机收集有用的杂交瘤来产生。为了证明这些单克隆抗体针对ATP酶亚基，我们测试了它们与纯化的豌豆子叶线粒体F1-ATP酶的交叉反应性。其中一种抗体（[α]-ATP酶D）与豌豆F1-ATP酶[α]-亚基交叉反应，两种抗体（[β]-ATP酶D和[β]-ATP酶E）与豌豆F1-ATP酶[β]-亚基交叉反应。这确定了，在9种抗体中，4种与玉米[α]-ATP酶亚基反应，另外5种与玉米[β]-ATP酶亚基反应。大多数单克隆抗体与来自广泛植物物种的F1-ATP酶交叉反应。针对[α]-亚基产生的四种单克隆抗体中的每一种识别不同的表位。在五种[β]-亚基抗体中，至少识别三种不同的表位。单克隆抗体与F1-ATP酶直接孵育不能抑制ATP酶活性。将单克隆抗体[α]-ATPaseD和[β]-ATPaseD与环氧玻璃QuantAffinity珠结合，并与纯化的豌豆F1-ATP酶制剂一起孵育。当抗体结合ATP酶时，ATP酶活性不受抑制。抗体被用来帮助豌豆F1-ATP酶亚基的整个豌豆子叶线粒体蛋白的二维地图。此外，这些抗体还揭示了在纯化的F1-ATP酶的[α]-和[β]-亚基中观察到的各种同种型之间的抗原相似性。这些单克隆抗体的特异性，沿着它们的跨物种识别和它们结合F1-ATP酶而不抑制酶功能的能力，使得这些抗体对于植物线粒体F1-ATP酶的进一步纯化和表征是有用的和无价的工具。
We have generated nine monoclonal antibodies against subunits of the maize (Zea mays L.) mitochondrial F1-ATPase. These monoclonal antibodies were generated by immunizing mice against maize mitochondrial fractions and randomly collecting useful hybridomas. To prove that these monoclonal antibodies were directed against ATPase subunits, we tested their cross-reactivity with purified F1-ATPase from pea cotyledon mitochondria. One of the antibodies ([alpha]-ATPaseD) cross-reacted with the pea F1-ATPase [alpha]-subunit and two ([beta]-ATPaseD and [beta]-ATPaseE) cross-reacted with the pea F1-ATPase [beta]-subunit. This established that, of the nine antibodies, four react with the maize [alpha]-ATPase subunit and the other five react with the maize [beta]-ATPase subunit. Most of the monoclonal antibodies cross-react with the F1-ATPase from a wide range of plant species. Each of the four monoclonal antibodies raised against the [alpha]-subunit recognizes a different epitope. Of the five [beta]-subunit antibodies, at least three different epitopes are recognized. Direct incubation of the monoclonal antibodies with the F1-ATPase failed to inhibit the ATPase activity. The monoclonal antibodies [alpha]-ATPaseD and [beta]-ATPaseD were bound to epoxide-glass QuantAffinity beads and incubated with a purified preparation of pea F1-ATPase. The ATPase activity was not inhibited when the antibodies bound the ATPase. The antibodies were used to help map the pea F1-ATPase subunits on a two-dimensional map of whole pea cotyledon mitochondrial protein. In addition, the antibodies have revealed antigenic similarities between various isoforms observed for the [alpha]- and [beta]-subunits of the purified F1-ATPase. The specificity of these monoclonal antibodies, along with their cross-species recognition and their ability to bind the F1-ATPase without inhibiting enzymic function, makes these antibodies useful and invaluable tools for the further purification and characterization of plant mitochondrial F1-ATPases.