PROPERTIES OF DOMINANT ANTIGENIC EPITOPES INFLUENCING MHC BINDING

MHC以抗原的形式呈递抗原的肽片段。 等位基因特异性的方式对T细胞。 从一个巨大的游泳池 加工过程中产生的潜在肽，仅选择一个子集 作为细胞表面的表位呈递，因此具有 可能引发T细胞反应。 选择的优势肽 由I-Ak从HEL衍生的氨基酸序列包含氨基酸HEL 48-62。 此前的数据显示， 提示HEL 48 -62表位的主要锚是D52。 这得到了实验的支持，在这些实验中， 残基显著降低了肽对 - Ak. 已经进行了一些初步实验， 鉴定A52 HEL的主要表位，并表明 48-62 A52肽不可检测。 44-61 A52已被 通过2-D免疫印迹鉴定为来自A52 HEL的主要I-Ak表位 HPLC和质谱法。 初始I-Ak竞争结合 实验表明，优势44-61 A52表艾德来自 A52 HEL对I-Ak的相对亲和力大于突变体48-62 A52肽。 44-61 A52肽的相对亲和力相等 与显性野生型加工的HEL 48-62肽相比。 在这 项目，我们正在努力开发的方法，以减少复杂性， 抗原肽混合物，通过使用2-D HPLC，第一维 第一种是离子交换色谱，第二种是毛细管RP-HPLC。

The MHC presents peptide fragments of antigens in an allele-specific manner to T cells. Out of an enormous pool of potential peptides generated during processing, only a select subset are presented as epitopes on the cell surface, and therefore have the potential to elicit T-cell responses. The dominant peptide selected from HEL by I-Ak comprises amino acids HEL 48-62. Previous data have suggested that the primary anchor of the HEL48-62 epitope is D52. This is supported by experiments in which alanine substitution of this residue drastically decreased the binding affinity of the peptide for I-Ak. A number of preliminary experiments have been performed to identify the major epitopes of A52 HEL and have indicated that the 48-62 A52 peptide is undetectable. Instead, 44-61 A52 has been identified as the predominant I-Ak epitope derived from A52 HEL by 2-D HPLC and mass spectrometry. Initial I-Ak competitive binding experiments show that the dominant 44-61 A52 epitope d eriv ed from A52HEL has a greater relative affinity for I-Ak than the mutant 48-62 A52 peptide. The relative affinity of the 44-61 A52 peptide is equal to that of the dominant wildtype processed HEL 48-62 peptide. In this project we are trying to develop methods to reduce the complexity of the antigenic peptide mixtures by using 2-D HPLC, the first dimension being ion-exchange chromatography and the second, capillary RP-HPLC.