Monoclonal antibodies specific for the empty conformation of HLA-DR1 reveal aspects of the conformational change associated with peptide binding
Monoclonal antibodies specific for the empty conformation of HLA-DR1 reveal aspects of the conformational change associated with peptide binding
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DOI：
10.1074/jbc.m314315200
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发表时间：
2004-04-16
影响因子：
4.8
通讯作者：
Stern, LJ
中科院分区：
文献类型：
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作者：
Carven, GJ;Chitta, S;Stern, LJ
第二类主要组织相容性复合体(MHC)蛋白结合多肽，并将它们呈现在细胞表面，与CD4(+)T细胞相互作用，作为免疫系统检查身体感染迹象的系统的一部分。基于多种流体力学和光谱学方法，已知多肽结合可以引起II类MHC蛋白的构象变化，但这种变化还没有明确地定位在整个II类MHC结构中。为了定位多肽诱导的人类常见的II类MHC变异体HLA-DR1的构象变化，我们产生了一系列识别空构象特异性的β亚基的单抗。对于在细胞表面表达的可溶性重组蛋白和天然蛋白，每个抗体都与空的形式反应，但不与多肽形式反应。抗体结合表位通过重叠多肽和丙氨酸扫描取代来表征，并定位于蛋白质的两个不同区域。表位内关键残基的模式表明，在肽结合过程中，这两个表位区域经历了实质性的构象变化。这些结果阐明了空形式的结构和多肽诱导的构象变化的本质。
Class II major histocompatibility complex (MHC) proteins bind peptides and present them at the cell surface for interaction with CD4(+) T cells as part of the system by which the immune system surveys the body for signs of infection. Peptide binding is known to induce conformational changes in class II MHC proteins on the basis of a variety of hydrodynamic and spectroscopic approaches, but the changes have not been clearly localized within the overall class II MHC structure. To map the peptide-induced conformational change for HLA-DR1, a common human class II MHC variant, we generated a series of monoclonal antibodies recognizing the beta subunit that are specific for the empty conformation. Each antibody reacted with the empty but not the peptide-loaded form, for both soluble recombinant protein and native protein expressed at the cell surface. Antibody binding epitopes were characterized using overlapping peptides and alanine scanning substitutions and were localized to two distinct regions of the protein. The pattern of key residues within the epitopes suggested that the two epitope regions undergo substantial conformational alteration during peptide binding. These results illuminate aspects of the structure of the empty forms and the nature of the peptide-induced conformational change.