Recombinant single chain human HLA-DR1 molecules as biologically active covalently linked heterotrimers of antigenic peptide, beta chain, and alpha chain

Xiaojie Zhu, Scheherazade Sadegh-Nasseri, Sina Bavari, Robert G. Ulrich, Michael Mage

Research output: Contribution to journalArticlepeer-review

Abstract

We engineered HLA-DR1 cDNA to encode influenza HA or HIV gag peptides covalently attached via a peptide linker to the amino terminus of the DR1 beta chain. Cotransfection with DR alpha cDNA into mouse L cells resulted in surface expression of HLA-DR1 molecules that reacted with mAbs specific for correctly folded HLA-DR epitopes. To ease production of these molecules, we engineered an additional peptide linker between the carboxyl terminus of a truncated beta chain (without TM or cytoplasmic domains) and the amino terminus of full length DR alpha chain. On transfection, the entire heterotrimer of peptide, beta chain, and alpha chain was expressed as a single covalently linked polypeptide chain with an expected molecular mass of approximately 66 kDa. Affinity purified soluble secreted single chain molecules (with truncated alpha chain) electrophorese as expected for compact Class II MHC dimers. Cell surface two-chain or single chain HLA-DR1 molecules with a covalent HA peptide can bind the SEA and SEB superantigens. They can also stimulate HLA-DR1 restricted HA-specific T cells. We plan to analyse the folding pathways of these recombinant MHC molecules and to study their in vitro immunogenicity.

Original languageEnglish (US)
Pages (from-to)A1175
JournalFASEB Journal
Volume10
Issue number6
StatePublished - 1996
Externally publishedYes

ASJC Scopus subject areas

  • Biotechnology
  • Biochemistry
  • Molecular Biology
  • Genetics

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