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 co...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 β2-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.展开更多
基金supported by Natural Science Fund of Guangdong Province (No.8451063201000340)the Talented Man Initiation Fund of Jinan University (No.51208004, No.51208017) to Dr. DY Ouyang +1 种基金grants from the National Natural Science Foundation of China (No.30572199, No.30230350 and No.30371651) to Prof. XH Hethe Biochemistry and Molecular Biology Key Discipline of Guangdong Province.
文摘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 β2-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.
