Cohen N, Crawford J S, Hiraki D D, Grumet F C
Department of Pathology, Stanford University School of Medicine, California.
Hum Immunol. 1989 Jul;25(3):207-222. doi: 10.1016/0198-8859(89)90083-9.
Two soluble, secreted forms of HLA-B7 were engineered by the creation of hybrid human/mouse molecules containing the polymorphic 5' region of the HLA-B7 gene and the secretory 3' region of the mouse Q10d gene. The hybrid, designated F1, is the first construct with only human extracytoplasmic domains, consisting of exons for the leader peptide and the three extracellular domains (alpha 1, alpha 2, alpha 3) of B7 spliced to the exons for the Q10d truncated transmembrane and 3' untranslated (3'UT) sequences. The second construct, designated C2, is similar but has the human alpha 3 replaced by the Q10 alpha 3 domain. Protein product from each construct was best demonstrated after gene transfection into the J27.2 cell line. In particular, secretion of the F1 product proves that the Q10 alpha 3 domain is not necessary for secretion of class I/Q10 hybrids. Moreover, the two soluble B7 forms, which differ only in their alpha 3 domain, are similarly recognized by monoclonal antibodies W6/32 (anti-HLA-ABC), BBM.1 (anti-human beta 2 microglobulin), and allo-B7-antibody, but differentially recognized by monoclonal antibody Q1/28 (anti-HLA class I heavy chain). Production of such soluble hybrid class I molecules in large amounts should allow critical structural and functional studies of these proteins.
通过构建包含HLA - B7基因多态性5'区域和小鼠Q10d基因分泌性3'区域的人/鼠杂交分子，设计出了两种可溶性、可分泌形式的HLA - B7。这种杂交分子命名为F1，是首个仅含人胞外结构域的构建体，由B7的前导肽外显子和三个胞外结构域（α1、α2、α3）与Q10d截短的跨膜和3'非翻译（3'UT）序列的外显子拼接而成。第二个构建体命名为C2，与之相似，但人α3结构域被Q10 α3结构域取代。将每个构建体的基因转染到J27.2细胞系后，能最好地展示出各构建体的蛋白质产物。特别地，F1产物的分泌证明Q10 α3结构域对于I类/Q10杂交分子的分泌并非必需。此外，这两种仅在α3结构域上存在差异的可溶性B7形式，同样能被单克隆抗体W6/32（抗HLA - ABC）、BBM.1（抗人β2微球蛋白）和同种异体B7抗体识别，但被单克隆抗体Q1/28（抗HLA I类重链）差异化识别。大量生产此类可溶性I类杂交分子应能对这些蛋白质进行关键的结构和功能研究。
