Kirchmair R, Leitner B, Fischer-Colbrie R, Marksteiner J, Hogue-Angeletti R, Winkler H
Department of Pharmacology, University of Innsbruck, Austria.
Biochem J. 1995 Aug 15;310 ( Pt 1)(Pt 1):331-6. doi: 10.1042/bj3100331.
We have established a radioimmunoassay for GE-25, a peptide present in the C-terminal end of the primary amino acid sequence of chromogranin A where it is flanked by typical proteolytic cleavage sites. Gel-filtration HPLC was used to characterize the molecular sizes of the immunoreactive molecules. The antiserum recognized not only the free peptide but also larger precursors including the proprotein chromogranin A. The tissues with the highest levels of GE-25 immunoreactivity were in decreasing order: the adrenal medulla, the three lobes of the pituitary gland, intestinal mucosa, pancreas and various brain regions. In adrenal medulla and parathyroid gland most of the immunoreactivity was found to be present as intact chromogranin A and some intermediate-sized peptides, without significant amounts of the free peptide. In anterior pituitary, and even more so in intestine, a shift to smaller peptides was seen. In the posterior and intermediate pituitary and in pancreas the predominant immunoreactive material was apparently represented by the free peptide GE-25. In reverse-phase chromatography this peptide eluted exactly like the synthetic standard, which allows a tentative identification as GE-25. In brain tissue the processing of chromogranin A was intermediate, with significant amounts of immunoreactivity corresponding to GE-25 as well as precursor proteins being present. We suggest that in those organs (endocrine pancreas, intermediate and posterior pituitary) where the major hormones are proteolytically processed there is also a concomitant proteolysis of further susceptible peptides. Since GE-25 is apparently formed in vivo and is well conserved between species it seems a good candidate for having specific physiological functions.
我们已经建立了一种针对GE - 25的放射免疫分析方法。GE - 25是一种肽,存在于嗜铬粒蛋白A一级氨基酸序列的C末端,其两侧为典型的蛋白水解切割位点。采用凝胶过滤高效液相色谱法来表征免疫反应性分子的分子大小。该抗血清不仅识别游离肽,还识别包括前体蛋白嗜铬粒蛋白A在内的更大的前体。GE - 25免疫反应性水平最高的组织按降序排列为:肾上腺髓质、垂体的三个叶、肠黏膜、胰腺和各个脑区。在肾上腺髓质和甲状旁腺中,大部分免疫反应性表现为完整的嗜铬粒蛋白A和一些中等大小的肽,游离肽含量不高。在垂体前叶,尤其是在肠道中,可见向较小肽的转变。在垂体后叶和中间叶以及胰腺中,主要的免疫反应性物质显然是游离肽GE - 25。在反相色谱中,该肽的洗脱情况与合成标准品完全一样,这使得可以初步鉴定为GE - 25。在脑组织中,嗜铬粒蛋白A的加工过程处于中间状态,存在大量与GE - 25以及前体蛋白相对应的免疫反应性。我们认为,在那些主要激素经过蛋白水解加工的器官(内分泌胰腺、垂体中间叶和后叶)中,也会伴随对其他易感肽的蛋白水解。由于GE - 25显然是在体内形成的,并且在物种间具有良好的保守性,它似乎是具有特定生理功能的良好候选者。