Protein Science, Vol 7, Issue 5 1245-1249, Copyright © 1998 by Cold Spring Harbor Laboratory Press

FOR THE RECORD

Assembly and crystallization of the complex between the human T cell coreceptor CD8{alpha} homodimer and HLA-A2

G. F. GAO, U. C. GERTH, J. R. WYER, B. E. WILLCOX, C. A. O'CALLAGHAN, Z. ZHANG, E. Y. JONES, J. I. BELL and B. K. JAKOBSEN
Molecular Immunology Group, Institute of Molecular Medicine, Oxford University, John Radcliffe Hospital, Oxford OX3 9DS, United Kingdom These authors contributed equally to this work.

A strategy for overexpression in Escherichia coli of the extracellular immunoglobulin domain of human CD8{alpha} was devised using codon usage alterations in the 5' region of the gene, designed so as to prevent the formation of secondary structures in the mRNA. A fragment of CD8{alpha}, comprising residues 1-120 of the mature protein, excluding the signal peptide and the membrane-proximal stalk region, was recovered from bacterial inclusion bodies and refolded to produce a single species of homodimeric, soluble receptor. HLA-A2 heavy chain, {beta}2-microglobulin and a synthetic peptide antigen corresponding to the pol epitope from HIV-1 were also expressed in E. coli, refolded and purified. CD8{alpha}/HLA-A2 complexes were formed in solution and by co-crystallization with a stoichiometry of one CD8{alpha}{alpha} dimer to one HLA-A2-peptide unit.
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