EFFICIENT MAPPING OF PROTEIN ANTIGENIC DETERMINANTS
EFFICIENT MAPPING OF PROTEIN ANTIGENIC DETERMINANTS
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DOI：
10.1073/pnas.83.18.7013
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发表时间：
1986-09-01
影响因子：
11.1
通讯作者：
YOUNG, RA
中科院分区：
文献类型：
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作者：
MEHRA, V;SWEETSER, D;YOUNG, RA
重组DNA表达策略已被用来推断麻风病病原体麻风分枝杆菌的单一蛋白质中六种不同抗原决定簇的氨基酸序列。对编码麻风分枝杆菌 65-kDa 抗原的基因进行了测序并得到了 lambda。 gt11基因子文库是用该基因的片段构建的。通过单克隆抗体筛选分离产生特异性抗原决定簇的重组DNA克隆，并通过快速引物延伸方法确定其插入DNA的序列。每个决定簇的氨基酸序列是根据多个抗体阳性DNA克隆的插入DNA的最小重叠推导出来的。阐明了六个不同表位的氨基酸序列。合成了含有这些表位之一的序列的肽，并显示其可以结合适当的单克隆抗体；这种抗原决定簇是麻风分枝杆菌所独有的。这里描述的方法可用于快速阐明抗体或 T 细胞识别的蛋白质表位。
A recombinant DNA expression strategy has been used to deduce the amino acid sequences of six different antigenic determinants in a single protein of Mycobacterium leprae, the etiologic agent of leprosy. The gene encoding the M. leprae 65-kDa antigen was sequenced and a .lambda. gt11 gene sublibrary was constructed with fragments of the gene. Recombinant DNA clones producing specific antigenic determinants were isolated by screening with monoclonal antibodies, and the sequences of their insert DNAs were determined with a rapid primer-extension method. The amino acid sequence of each determinant was deduced from the minimum overlap of insert DNAs from multiple antibody-positive DNA clones. Amino acid sequences for six different epitopes were elucidated. A peptide containing sequences for one of these epitopes was synthesized and shown to bind the appropriate monoclonal antibody; this antigenic determinant is unique to M. leprae. The approach described here can be used to rapidly elucidate protein epitopes that are recognized by antibodies or T cells.