Characterisation of Zea mays L. plastidial transglutaminase: interactions with thylakoid membrane proteins
Characterisation of Zea mays L. plastidial transglutaminase: interactions with thylakoid membrane proteins
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DOI：
10.1111/j.1438-8677.2009.00280.x
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发表时间：
2010-09-01
期刊：
影响因子：
3.9
通讯作者：
Santos, M.
中科院分区：
文献类型：
--
作者：
Campos, A.;Carvajal-Vallejos, P. K.;Santos, M.
叶绿体转氨酶（chlTGase）活性在植物对光刺激的响应和光适应中起重要作用，但其在叶绿体中的确切功能尚不清楚。在蛋白质组学水平上，chlTGase与类囊体蛋白的相互作用和其与光系统II（PSII）蛋白复合物的关联的演示的表征与实验使用玉米类囊体蛋白提取物完成。通过针对玉米TGase基因产物的C-末端序列设计的特异性抗体，在玉米叶绿体分化的三个不同阶段的类囊体膜提取物中免疫检测到不同的chlTGase形式。这些条带与lhcb 1、2和3触角蛋白的条带共定位。最重要的是，一个58 kDa的形式存在于成熟的叶绿体，其特征在于使用生物化学和蛋白质组学方法。从光诱导成熟叶绿体类囊体蛋白的连续分级显示，58 kDa的形式与类囊体膜，表现为可溶性或外周膜蛋白。二维凝胶电泳区分，第一次，58 kDa的带在两种不同的形式，可能对应于两个不同的TGase cDNA先前克隆。电泳分离天然凝胶中的类囊体蛋白，然后通过LC-MS质谱鉴定蛋白复合物表明，玉米chlTGase形成一个特定的PSII蛋白复合物，其中包括LHCII，ATP酶和pSbS蛋白的一部分。结果进行了讨论，这些蛋白质之间的相互作用和建议的作用的酶在类囊体膜组织和光保护。
Chloroplast transglutaminase (chlTGase) activity is considered to play a significant role in response to a light stimulus and photo-adaptation of plants, but its precise function in the chloroplast is unclear. The characterisation, at the proteomic level, of the chlTGase interaction with thylakoid proteins and demonstration of its association with photosystem II (PSII) protein complexes was accomplished with experiments using maize thylakoid protein extracts. By means of a specific antibody designed against the C-terminal sequence of the maize TGase gene product, different chlTGase forms were immunodetected in thylakoid membrane extracts from three different stages of maize chloroplast differentiation. These bands co-localised with those of lhcb 1, 2 and 3 antenna proteins. The most significant, a 58 kDa form present in mature chloroplasts, was characterised using biochemical and proteomic approaches. Sequential fractionation of thylakoid proteins from light-induced mature chloroplasts showed that the 58 kDa form was associated with the thylakoid membrane, behaving as a soluble or peripheral membrane protein. Two-dimensional gel electrophoresis discriminated, for the first time, the 58-kDa band in two different forms, probably corresponding to the two different TGase cDNAs previously cloned. Electrophoretic separation of thylakoid proteins in native gels, followed by LC-MS mass spectrometry identification of protein complexes indicated that maize chlTGase forms part of a specific PSII protein complex, which includes LHCII, ATPase and pSbS proteins. The results are discussed in relation to the interaction between these proteins and the suggested role of the enzyme in thylakoid membrane organisation and photoprotection.