Preliminary X-ray diffraction analysis of crystals from the recombinantly expressed human major histocompatibility antigen HLA-B*2704 in complex with a viral peptide and with a self-peptide

Abstract

The product of the human leukocyte antigen (HLA) gene HLA-B*2704 differs from that of the prototypical subtype HLA-B*2705 by three amino acids at heavy-chain residues 77 (Ser instead of Asp), 152 (Glu instead of Val) and 211 (Gly instead of Ala). In contrast to the ubiquitous HLA-B*2705 subtype, HLA-B*2704 occurs only in orientals. Both subtypes are strongly associated with spondyloarthropathies and the peptides presented by these subtypes are suspected to play a role in disease pathogenesis. HLA-B*2704 was crystallized in complex with a viral peptide and with a self-peptide using the hanging-drop vapour-diffusion method with PEG as a precipitant. Both crystals belong to space group P2(1)2(1)2(1). Data sets were collected to 1.60 A (complex with the self-peptide pVIPR) or to 1.90 A (complex with the viral peptide pLMP2) resolution using synchrotron radiation. With HLA-B*2705 complexed with pVIPR as a search model, unambiguous molecular-replacement solutions were found for the complexes of HLA-B*2704 with both peptides.

Figure 1

Analysis of the purity of the refolded B*2704 complexes. Samples were subjected to SDS–PAGE under reducing (lanes 2 and 4) or non-reducing (lanes 3 and 5) conditions and stained with Coomassie Brilliant Blue. Lane 1, molecular-weight markers (kDa), lanes 2 and 3, B*2704–pLMP2; lanes 4 and 5, B*2704–pVIPR. The positions of HLA-B27 HC and β₂m are indicated. The impurity with slightly higher molecular weight than the HC is occasionally found in preparations of HLA class I molecules. We have so far not observed it to interfere with the crystallization of the complexes.

Figure 2

Crystals of B*2704–pLMP2. The black bar indicates a length of 50 µm. Crystals of B*2704–pVIPR exhibited comparable size and morphology.