Gill B M, Barbosa J A, Dinh T Q, Garrod S, O'Connor D T
Department of Medicine, University of California, San Diego.
Regul Pept. 1991 Apr 25;33(2):223-35. doi: 10.1016/0167-0115(91)90216-4.
The chromogranins/secretogranins are a family of neuroendocrine vesicle secretory proteins. Immunohistology and immunoblotting have suggested that a major soluble protein in human chromaffin granules may be chromogranin B (CgB). We purified from pheochromocytoma chromaffin granules an SDS-PAGE 110-120 kDa protein whose N-terminal sequence matched that previously deduced from a human CgB cDNA. An antibody directed against a synthetic human CgB N-terminal region specifically recognized the CgB N-terminus, though not the chromogranin A (CgA) N-terminus or the CgB C-terminus on immunoblots. An antiserum directed against CgB's C-terminus also visualized CgB but not CgA. By immunoblotting, CgB was a quantitatively major protein in human pheochromocytoma chromaffin granules, but a relatively minor in normal bovine adrenal medullary chromaffin granules. In a variety of normal bovine neuroendocrine tissues, the relative abundance of CgB immunoreactivity on immunoblots was: adrenal medulla greater than anterior pituitary greater than pancreas greater than small intestine, hypothalamus. Immunoblotting of neuroendocrine tissues (or their hormone storage vesicle cores) with both anti N-terminal and anti C-terminal CgB antisera suggested bidirectional cleavage or processing of CgB; in the anterior pituitary, a unique 40 kDa C-terminal fragment was observed. Bidirectional CgB cleavage was also suggested on immunoblots of chromaffin tissue from three species (human, bovine, rat). C-terminal processing of CgB was also confirmed by amino acid sequencing of SDS-PAGE-separated, polyvinylidene difluoride membrane-immobilized CgB fragments from pheochromocytoma chromaffin granules. Whether such fragments possess biological activity remains to be investigated.
嗜铬粒蛋白/分泌粒蛋白是一类神经内分泌囊泡分泌蛋白。免疫组织学和免疫印迹表明，人嗜铬粒中的一种主要可溶性蛋白可能是嗜铬粒蛋白B（CgB）。我们从嗜铬细胞瘤嗜铬粒中纯化出一种SDS-PAGE 110 - 120 kDa的蛋白，其N端序列与先前从人CgB cDNA推导的序列相符。一种针对合成人CgB N端区域的抗体在免疫印迹上能特异性识别CgB的N端，但不能识别嗜铬粒蛋白A（CgA）的N端或CgB的C端。一种针对CgB C端的抗血清也能检测到CgB，但检测不到CgA。通过免疫印迹分析，CgB是人类嗜铬细胞瘤嗜铬粒中的主要定量蛋白，但在正常牛肾上腺髓质嗜铬粒中相对较少。在多种正常牛神经内分泌组织中，免疫印迹上CgB免疫反应性的相对丰度为：肾上腺髓质大于垂体前叶大于胰腺大于小肠、下丘脑。用抗CgB N端和抗CgB C端抗血清对神经内分泌组织（或其激素储存囊泡核心）进行免疫印迹分析，提示CgB存在双向切割或加工；在垂体前叶，观察到一个独特的40 kDa C端片段。来自三种物种（人、牛、大鼠）的嗜铬组织免疫印迹也提示了CgB的双向切割。通过对嗜铬细胞瘤嗜铬粒中SDS-PAGE分离、固定在聚偏二氟乙烯膜上的CgB片段进行氨基酸测序，也证实了CgB的C端加工。这些片段是否具有生物活性仍有待研究。
