Construction of Soluble Mamu-B*1703, a Class I Major Histocompatibility Complex of Chinese Rhesus Macaques, Monomer and Tetramer Loaded with a Simian Immunodeficiency Virus Peptide
Construction of Soluble Mamu-B*1703, a Class I Major Histocompatibility Complex of Chinese Rhesus Macaques, Monomer and Tetramer Loaded with a Simian Immunodeficiency Virus Peptide
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DOI：
10.1038/cmi.2009.16
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发表时间：
2009-04-01
影响因子：
24.1
通讯作者：
Guo, He
中科院分区：
文献类型：
--
作者：
Ouyang, Dongyun;Wang, Xiaoying;Guo, He
在HIV感染和疫苗研究中，华裔猕猴比印度血统的猕猴更普遍。然而，目前可用的大多数数据和试剂，如主要组织相容性复合体(MHC)I类四聚体，都来自印度起源的猕猴，因为这些动物在历史上占主导地位。虽然这两个猕猴群体的免疫遗传背景有显著差异，但它们有一些共同的MHC I类等位基因。本研究报道了携带猴免疫缺陷病毒(SIV)优势表位IW9(IRYPKTFGW)的可溶性MAMU-B*1703(中国猕猴MHC类分子)单体和四聚体的制备方法，该表位在印度猕猴中被鉴定为MAMU-B*1701限制性的。从已克隆的MAMU-B*1703胞外区与BIRA底物多肽(BSP)融合的DNA片段中，扩增出MAMU-B*1703胞外区的DNA片段，并将其插入原核表达载体。在抗原肽IW9和轻链β(2)-微球蛋白存在下，表达的重链被复性为可溶的单体。生物素化后，四个单体被藻红蛋白偶联的链霉亲和素聚合成四聚体。该四聚体已被证实具有正确的构象，是研究中国猕猴SIV疫苗模型中抗原特异性CD8(+)T淋巴细胞的潜在工具。我们的结果还表明，在印度猕猴中报道的一些抗原肽可以直接作为构建中国猕猴MHC四聚体的配基。细胞与分子免疫学。2009年；6(2)：117-122。
Chinese-descent rhesus macaques have become more prevalent for HIV infection and vaccine investigation than Indian-origin macaques. Most of the currently available data and reagents such as major histocompatibility complex (MHC) class I tetramers, however, were derived from Indian-origin macaques due to the dominant use of these animals in history. Although there are significant differences in the immunogenetic background between the two macaque populations, they share a few of common MHC class I alleles. We reported in this study the procedure for preparation of a soluble Mamu-B*1703 (a MHC class I molecule of Chinese macaques) monomer and tetramer loaded with a dominant simian immunodeficiency virus (SIV) epitope IW9 (IRYPKTFGW) that was identified to be Mamu-B*1701-restricted in Indian macaques. The DNA fragment encoding the Mamu-B*1703 extracellular domain fused with a BirA substrate peptide (BSP) was amplified from a previously cloned cDNA and inserted into a prokaratic expression vector. In the presence of the antigenic peptide IW9 and light chain beta(2)-microglobulin, the expressed heavy chain was refolded into a soluble monomer. After biotinylation, four monomers were polymerized as a tetramer by phycoerythrin-conjugated streptavidin. The tetramer, having been confirmed to have the right conformation, was a potential tool for investigation of antigen-specific CD8(+) T-lymphocytes in SIV vaccine models of Chinese macaques. And our results also suggested that some antigenic peptides reported in Indian-origin macaques could be directly recruited as ligands for construction of Chinese macaque MHC tetramers. Cellular & Molecular Immunology. 2009;6(2):117-122.