Regulatory mechanism of olfactory reception in olfactory cells

在这项研究中，我们从青蛙嗅觉上皮细胞中分离到一种新的26 kDa钙结合蛋白p26olf，除了凝胶过滤外，还经过了苯基琼脂糖层析、DEAE琼脂糖层析和Mono-Q层析。提取的蛋白用赖氨酰内切酶消化，得到p26olf的蛋白水解物。根据蛋白质测序仪分析的氨基酸序列，我们合成了DNA探针，并利用青蛙嗅觉上皮的cDNA文库进行了聚合酶链式反应。获得了一个全长806个碱基的cDNA克隆。该基因推导的氨基酸序列包含了多肽片段中的所有氨基酸序列。因此，我们得出结论，该基因编码p26olf。结果表明，p26olf由217个氨基酸组成，分子量为24,493 kDa。同源性搜索分析表明，p26olf的N-末端与牛S-100β的氨基酸序列有34.1%的同源性，C-末端的氨基酸序列与牛S-100α的氨基酸序列有32.3%的同源性。由于S-100蛋白具有二聚体的功能，p26olf的功能可能与S-100蛋白相似，只是以单体形式存在。Northern印迹分析表明，p26olf在青蛙嗅觉上皮细胞以及肺、脾等其他组织中均有表达。在嗅上皮的纤毛层中发现了一种针对p26olf多肽的抗体，这些结果表明p26olf是S-100蛋白的二聚体，参与了嗅觉转导或适应。

In this study, we isolated a novel 26 kDa Ca-binding protein named p26olf from frog olfactory epithelium after several chromatographycal steps including phenyl Sepharose, DEAE Sepharose and Mono Q chromatography in addition to gel filtration. The isolated protein was digested with lysyl endopeptidase to obtain proteolytic fragments of p26olf. Based on the amino acid sequence of the fragments analyzed by a protein sequencer, we synthesized a DNA probe and did PCR using cDNA library of the frog olfactory epithelium. We obtained a cDNA clone consisting of 806 bp. Deduced amino acid sequence of this cDNA contained all of the amino acid sequences identified in the peptide fragments. For this reason, we concluded that this cDNA encodes p26olf. It was found that p26olf consists of 217 amino acids with a calculated molecular mass of 24,493 kDa. Homology search analysis showed that the amino acid sequence of the N-terminal half of p26olf exhibits 34.1 % identity to that of bovine S-100beta and the C-terminal half 32.3 % identity to that of bovine S-100 alpha. Since S-100 proteins are functional as form of dimer, p26olf may function similarly as S-100 proteins but in the form of monomer. Northern blot analysis showed that p26olf is expressed in the frog olfactory epithelium as well as in other tissues like lung and spleen. Immunoreactivity of an antibody raised against a p26olf peptide was found in the cilia layr of the olfactory epithelium.Taken together, these results suggest that p26olf is a dimeric form of S-100 protein and is involved in olfactory transduction or adaptation.