A Novel Protein from Germinating Pollen with OxO activity; Characterization and Functional Testing

一种来自发芽花粉的具有 OxO 活性的新型蛋白质；

• 批准号: 0216964
• 负责人: Loverine Taylor
• 金额: --
• 依托单位: Washington State University
• 依托单位国家: 美国
• 项目类别: Standard Grant
• 财政年份: 2002
• 资助国家: 美国
• 起止时间: 2002-09-01 至 2004-08-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=0216964&HistoricalAwards=false
• 关键词: Novel; Protein; Germinating; Pollen; OxO

Abstract0216964 Taylor Pollen tubes deliver sperm to the ovules via a process known as tip growth. Large amounts of plasma membrane and cell wall precursors, synthesized in the Golgi, are transported to the tip where they undergo the polymerization and cross-linking reactions characteristic of tip growth. This proposal describes a pollen-specific protein, CBP25/22, that was isolated by a functional interaction with calmodulin (CaM), the major Ca 2+-sensing protein in cells. CBP25/22 is located on the exterior face of an abundant class of pollen organelles and has oxalate oxidase (OxO) activity, a reaction that generates peroxide (H2O2). Based on these unique biochemical properties, we propose that the CBP25/22 OxO-generated H2O2 mediates cross-linking of wall proteins and polymers in growing pollen tubes. Moreover, given the pivotal role of tip Ca2+ gradients in pollen tube growth, we propose that the Ca 2+-sensitive binding of CaM to CBP25/22 confines expression of the OxO activity to the growing tip. The focus of this research is to demonstrate that the unique biochemical properties of CBP25/22 identified in vitro, are expressed within the growing pollen tube. The proposed experimental plan will use immunological, biochemical and protein-protein interaction techniques to localize the CBP25/22 and CaM proteins and the enzymatic product (H2O2 ) in situ . Specifically we will demonstrate a Ca 2+ -sensitive association of CBP25/22 and CaM in vivo and show that OxO-generated H2O2 co-localizes with the CBP25/22 protein in vivo. This research may provide a novel way to control pollination, thus alleviating the current concern about gene flow via pollen from transgenic crops. In addition it may have a broader impact on basic growth mechanisms; neurons and fungal hypha also exhibit tip growth. Significantly, CBP25/22 shares its unique CaM-binding characteristics with neuromodulin, a protein that functions in vesicle transport and fusion in the tip of extending neurons. This similarity suggests that there may be a common mechanism underlying tip growth in diverse organisms.

泰勒花粉管通过一个称为尖端生长的过程将精子输送到胚珠。在高尔基体中合成的大量质膜和细胞壁前体被运输到尖端，在那里它们经历尖端生长特征的聚合和交联反应。该建议描述了一种花粉特异性蛋白质，CBP 25/22，其通过与细胞中主要的Ca 2+敏感蛋白钙调素（CaM）的功能相互作用而分离。CBP 25/22位于丰富的花粉细胞器的外表面，具有草酸氧化酶（OxO）活性，这是一种产生过氧化物（H2 O2）的反应。基于这些独特的生物化学性质，我们提出，CBP 25/22 OxO产生的H2 O2介导的交联壁蛋白和聚合物在生长的花粉管。此外，由于顶端Ca 2+梯度在花粉管生长中的关键作用，我们提出CaM与CBP 25/22的Ca 2+敏感结合限制了生长顶端的OxO活性的表达。本研究的重点是证明在体外鉴定的CBP 25/22的独特生化特性在生长的花粉管中表达。拟议的实验计划将使用免疫学、生物化学和蛋白质-蛋白质相互作用技术来原位定位CBP 25/22和CaM蛋白质以及酶产物（H2 O2）。具体来说，我们将证明在体内的CBP 25/22和钙调素的钙2+敏感的协会，并显示OxO产生的H2 O2与CBP 25/22蛋白在体内共定位。这项研究可能提供一种控制授粉的新方法，从而缓解目前人们对转基因作物花粉基因流动的担忧。此外，它可能对基本生长机制产生更广泛的影响;神经元和真菌菌丝也表现出尖端生长。值得注意的是，CBP 25/22与神经调节素（一种在延伸神经元的尖端中的囊泡运输和融合中起作用的蛋白质）共享其独特的CaM结合特征。这种相似性表明，在不同的生物体中，尖端生长可能有一个共同的机制。