NATURE OF PEPTIDE BINDING IN HLA-A*66 ALLELIC VARIANTS.

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NATURE OF PEPTIDE BINDING IN HLA-A*66 ALLELIC VARIANTS.
Christina Bade-Doeding , Britta Eiz-Vesper PhD , Holger-A. Elsner MD, PhD , Axel Seltsam MD , Joachim Kuhn PhD and Rainer Blasczyk MD, PhD . Hannover Germany, Hannover Medical School, D-30625, Department of Transfusion Medicine .

HLA class I molecules present peptides which are usually 8-10 amino acids in length, and which essentially contribute to the antigenicity of the molecule. Few studies have addressed the question to which extent variants of the same allelic group differ in their peptide motifs. To answer this question for the HLA-A*66 group, we have sequenced the eluate of a recombinant A*6602 molecule, and compared it with peptide sequence data reported for A*6601. Both A*66 variants differ by position 90 (D→A), located in an outer loop, and position 163 (R→E) shaping pockets A and B. The truncated recombinant product of A*6602 was expressed in the HLA class I deficient cell line LCL 721.221. Cell culture supernatant was purified by W6/32 affinity chromatography, peptides were obtained by acid elution, and separated by RP-HPLC. Sequences were determined by MALDI-TOF mass spectrometry. For A*6601, at position 2 of the bound peptides, T and V; and at position 9, R have been previously described as anchor residues, bound by pockets B and F, respectively. In A*6602 derived peptides, the same anchors at position 9 as in A*6601 peptides were found, reflecting their identical structure with regard to pocket F. Moreover, also position 2 corresponded to the anchor residues reported for A*6601, although the pocket B shaping position 163 has been described as critical regarding HLA class I allorecognition. This might be explained by the observation that R and E frequently replace each other without affecting protein function. These results suggest that in A*66 the R→E exchange leaves the P2 anchor of the peptide unaffected. Accordingly, this variation appears not to alter significantly the binding characteristics of the A*66 pockets allowing to expect a low allogenicity between A*6601 and A*6602.