Secretion of genetically engineered human/mouse class I antigens.
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.
Cohen
N
N
Department of Pathology, Stanford University School of Medicine, California.
Crawford
J S
JS
Hiraki
D D
DD
Grumet
F C
FC
D000818:Animals; D000911:Antibodies, Monoclonal; D002460:Cell Line; D002469:Cell Separation; D004591:Electrophoresis, Polyacrylamide Gel; D004797:Enzyme-Linked Immunosorbent Assay; D005434:Flow Cytometry; D005455:Fluorescent Antibody Technique; D005818:Genetic Engineering / Q000379:methods*; D015395:Histocompatibility Antigens Class I / Q000378:metabolism*; D006801:Humans; D006825:Hybridomas / Q000378:metabolism*; D051379:Mice; D011233:Precipitin Tests; D014162:Transfection
D000911:Antibodies, Monoclonal; D015395:Histocompatibility Antigens Class I