D-B-BINDING PEPTIDES FROM INFLUENZA-VIRUS - EFFECT OF NON-ANCHOR RESIDUES ON STABILITY AND IMMUNODOMINANCE

Relative affinities were determined for the interaction of H-2D(b) wit h all the peptides from the A/PR/8/34 strain of influenza virus that c ontained the Db-binding motif. The results indicated that, even though 23 peptides with the appropriate motif were identified and analysed, binding of only five of them could be detected at peptide concentratio ns lower than 10(-7) M. Of these five, only one, TGICNQNII, bound with better affinity than the nucleoprotein-derived natural epitope, ASNEN METM. The origin of the higher binding peptide was the influenza neura minidase, a protein for which little cytosolic processing would be exp ected since it is a surface glycoprotein. To establish why many of the influenza-derived peptides did not bind, the role of non-anchor resid ues on Db-peptide interactions was analysed, using a scheme where QDIE NEEKI, a non-binding peptide from the influenza virus polymerase 1, wa s sequentially converted to ASNENMETI, which binds to Db With an affin ity similar to that of ASNENMETM. Although all positions examined infl uenced peptide binding, peptide residue no. 2 (P2) was of particular i mportance. Therefore, each of the 20 naturally occurring amino acids w ere inserted at this position to investigate their effects on peptide- MHC interaction. The results indicated that amino acids having side ch ains with charged or ring structures were deleterious, while non-polar and polar residues were either neutral or facilitated binding to diff erent degrees. Our data also indicated that every residue of the pepti de contributes to the stability of the MHC-peptide complex, and the fi nal affinity is dependent on the nature of the amino acids at each pos ition, not just on those at a small number of anchor positions. The re sults also suggested that increased stability, as indicated by the hal f-life of the peptide-MHC class I complex, might play an important rol e in selecting the immunodominant epitope.