Purification, structural characterization, cloning and immunocytochemical localization of chemoreception proteins from Schistocerca gregaria
Purification, structural characterization, cloning and immunocytochemical localization of chemoreception proteins from Schistocerca gregaria
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DOI：
10.1046/j.1432-1327.1999.00438.x
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发表时间：
1999-06-01
期刊：
影响因子：
--
通讯作者：
Pelosi, P
中科院分区：
文献类型：
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作者：
Angeli, S;Ceron, F;Pelosi, P
从沙漠蝗Schistocerca gregaria的化学感受器官（触角，跗关节和唇）中纯化了可溶性低分子量蛋白异构体。从跗关节末端cDNA序列中PCR扩增出5个编码这类蛋白质的基因。它们的表达产物是109个氨基酸的多肽链，与来自黑腹果蝇和Cactoblastis cactorum的推定嗅觉蛋白具有40-50%的序列同一性。从化学感受器官纯化的异构体的直接结构研究揭示了克隆的两个基因的表达产物中的存在。另外两种蛋白质亚型被检测到，其分子结构详尽的特点。所有亚型的MS分析表明，多肽链中保守的4个半胱氨酸残基参与二硫键（Cys 29-Cys 38和Cys 57-Cys 60），并表明不存在任何额外的翻译后修饰。免疫细胞化学实验，进行兔抗血清提出的蛋白质异构体的混合物，显示选择性标记的外淋巴接触感器的跗骨，下颚须和触角。其他类型的感器没有标记，也没有感觉细胞的角质层和树突。未检测到放射性标记的葡萄糖或碳酸氢盐的结合，与这类蛋白质参与CO2传感级联的假设不一致。我们的实验数据表明，这里描述的蛋白质可能参与直翅目接触化学感受。
Soluble low-molecular-mass protein isoforms were purified from chemosensory organs (antennae, tarsi and labrum) of the desert locust Schistocerca gregaria. Five genes encoding proteins of this group were amplified by PCR from cDNAs of tarsi end sequenced. Their expression products are polypeptide chains of 109 amino acids shelling 40-50% sequence identity with putative olfactory proteins from Drosophila melanogaster and Cactoblastis cactorum. Direct structural investigation on isoforms purified from chemosensory organs revealed the presence in the expression products of two of the genes cloned. Two additional protein isoforms a ere detected and their molecular structure exhaustively characterized. MS analysis of all isoforms demonstrated that the four cysteine residues conserved in the polypeptide chain were involved in disulfide bridges (Cys29-Cys38 and Cys57-Cys60) and indicated the absence of any additional post-translational modifications. Immunocytochemistry experiments, performed with rabbit antiserum raised against the protein isoform mixture, showed selective labelling of the outer lymph in contact sensilla of tarsi, maxillary palps and antennae. Other types of sensilla were not labelled, nor were the cuticle and dendrites of the sensory cells. No binding of radioactively labelled glucose or bicarbonate was detected, in disagreement with the hypothesis that this class of proteins is involved in the CO2-sensing cascade. Our experimental data suggest that the proteins described here could be involved in contact chemoreception in Orthoptera.