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Volume 271, Number 33, Issue of August 16, 1996 pp. 20156-20162
©1996 by The American Society for Biochemistry and Molecular Biology, Inc.

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Expression of Recombinant HLA-DR2 Molecules
REPLACEMENT OF THE HYDROPHOBIC TRANSMEMBRANE REGION BY A LEUCINE ZIPPER DIMERIZATION MOTIF ALLOWS THE ASSEMBLY AND SECRETION OF SOLUBLE DR HETERODIMERS

(Received for publication, March 18, 1996)

Avtandil Kalandadze , Michael Galleno ^§ , Luis Foncerrada ^§ , Jack L. Strominger and Kai W. Wucherpfennig ^¶

From the  Dana-Farber Cancer Institute and ^¶ Department of Neurology, Harvard Medical School, Boston, Massachusetts 02115 and the ^§ Invitrogen Corporation, San Diego, California 92121

Major histocompatibility complex (MHC) class II molecules are membrane-anchored heterodimers that present peptides on the surface of antigen presenting cells to T cells. Soluble HLA-DR2 molecules were expressed for structural and functional characterization of the MHC/peptide/T cell receptor recognition unit. The  and  chains of DR2 (encoded by the DRA, DRB1*1501 genes) did not assemble in mammalian or insect cell lines when the transmembrane regions of one or both chains were truncated. The hydrophobic transmembrane regions of DR and DR facilitate assembly of the heterodimer and were therefore replaced by the leucine zipper dimerization motifs from the transcription factors Fos and Jun, which assemble as a soluble, tightly packed coiled coil structure. The DR-Fos and DR-Jun constructs were expressed in a methyltrophic yeast, Pichia pastoris, using the -mating factor secretion signal to direct expression to the secretory pathway. DR heterodimers were purified from supernatants using an antibody specific for the DR heterodimer. Kinetic and quantitative peptide binding experiments demonstrated that recombinant DR2 molecules were efficiently loaded with an antigenic peptide. Soluble DR2 molecules can be used to define structural aspects of the MHC/peptide/T cell receptor interaction and to study the signals induced by T cell receptor recognition of soluble DR2·peptide complexes.

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