MOLECULAR TOPOGRAPHY OF THE NEURAL CELL-ADHESION MOLECULE N-CAM - SURFACE ORIENTATION AND LOCATION OF SIALIC ACID-RICH AND BINDING REGIONS
MOLECULAR TOPOGRAPHY OF THE NEURAL CELL-ADHESION MOLECULE N-CAM - SURFACE ORIENTATION AND LOCATION OF SIALIC ACID-RICH AND BINDING REGIONS
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DOI：
10.1073/pnas.80.10.3116
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发表时间：
1983-01-01
期刊：
影响因子：
--
通讯作者：
EDELMAN, GM
中科院分区：
文献类型：
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作者：
CUNNINGHAM, BA;HOFFMAN, S;EDELMAN, GM
为阐明鸡胚脑神经细胞粘附分子（N-CAM）的结构与功能关系，进行了化学分析和结合研究。从细胞表面分离的N-CAM似乎包括2条密切相关的多肽链。用神经氨酸酶处理这种与固体载体上的抗体结合的N-CAM制剂，得到MW为140，000和170，000的组分。这些组分各自具有与N-CAM相同的氨基末端序列，并且在肽图谱上给出了几乎相同的图谱。来自用衣霉素处理的9天脑细胞的N-CAM的免疫沉淀产生MW为130，000和160，000的相应组分，表明N-CAM的这两种组分之间的差异在于多肽而不是分子的碳水化合物部分。N-CAM的氨基末端似乎远离细胞表面延伸，大部分唾液酸靠近肽链中部。在37 ℃下孵育N-CAM。C产生MW 65，000的片段（Fr 1），其缺乏大部分唾液酸。用金黄色葡萄球菌V-8蛋白酶处理膜释放出含有大部分唾液酸的N-CAM片段（Fr 2）;神经氨酸酶处理后，该片段的MW为108，000。这两个片段都含有多肽链的氨基末端部分。N-CAM结合位点的至少一部分位于肽链的氨基末端区域。大多数或所有的唾液酸不直接参与结合，尽管它可以影响结合，如通过发现神经氨酸酶处理的N-CAM（去唾液酸化-N-CAM）比未处理的N-CAM更大程度地结合细胞所指示的。溶液中的Fr 1和Fr 2片段不与细胞结合，但作为N-CAM与细胞结合的竞争者，与N-CAM和去唾液酸化-N-CAM一样有效。当共价固定到珠子上时，N-CAM、去唾液酸化-N-CAM和Fr 1和Fr 2片段特异性地结合到细胞上。N-CAM自溶产物与Fr 1一起沿着释放，既不与细胞结合，也不与N-CAM竞争结合。除了建议的N-CAM结合区域的位置，积累的结果提高的可能性，价可能在N-CAM结合中发挥关键作用。
Chemical analyses and binding studies were correlated to clarify the relationship of structure to function in the neural cell adhesion molecule (N-CAM) from embryonic chicken brain. N-CAM isolated from the cell surface appears to include 2 closely related polypeptide chains. Treatment with neuraminidase of such preparations of N-CAM bound by antibodies on solid supports yielded components of MW 140,000 and 170,000. These components each had the same amino-terminal sequence as N-CAM and gave nearly identical profiles on peptide maps. Immunoprecipitation of N-CAM from 9-day brain cells treated with tunicamycin yielded corresponding components of MW 130,000 and 160,000, suggesting that the differences between these 2 components of N-CAM are in the polypeptide rather than the carbohydrate portions of the molecules. N-CAM appears to be oriented with the amino terminus extending away from the cell surface and with the bulk of the sialic acid near the middle of the peptide chain. Incubation of N-CAM at 37.degree. C generates a fragment (Fr1) of MW 65,000 that lacks most of the sialic acid. Treatment of membranes with Staphylococcus aureus V-8 protease released a fragment (Fr2) of N-CAM that contained most of the sialic acid; this fragment had an MW of 108,000 after neuraminidase treatment. Both of these fragments contain the amino-terminal portion of the polypeptide chain. At least a portion of the N-CAM binding site was located in the amino-terminal region of the peptide chain. Most or all of the sialic acid was not directly involved in binding, although it can influence binding, as indicated by the finding that neuraminidase-treated N-CAM (desialylated-N-CAM) bound to cells to a greater extent than untreated N-CAM. The Fr1 and the Fr2 fragments in solution did not bind to cells but were as effective as N-CAM and desialylated-N-CAM as competitors for N-CAM binding to cells. When fixed covalently to beads, N-CAM, desialylated-N-CAM and Fr1 and Fr2 fragments bound specifically to cells. The N-CAM autolysis products released along with Fr1 neither bound to cells nor competed for N-CAM binding. In addition to suggesting a location for the N-CAM binding region, the accumulated results raise the possibility that valence may play a key role in N-CAM binding.