| Abstract: | Laser Raman spectroscopy has been employed to study the structure of the hen egg yolk protein phosvitin in H2O and D2O solutions at neutral and acidic pH (pD) and in the solid state. The Raman data indicate an unusual conformation for phosvitin in neutral aqueous solution, which is deficient in both alpha-helix and conventional beta-sheet conformations. This unusual pH 7 structure is, however, largely converted to a beta-sheet conformation in strongly acidic media (pH less than 2). beta-Sheet is also the predominant secondary structure for phosvitin in the solid state, obtained by lyophilization of the protein from aqueous solution at neutral pH. The imidazolium rings of histidyl residues remain significantly protonated near neutrality, which suggests substantial elevation of the pK for imidazolium ring ionizations of phosvitin in aqueous solution. This may result from extensive ion-pair interactions involving positively charged histidines and negatively charged phosphoserines, which are prevalent in the phosvitin sequence. The present results suggest that antiparallel beta-sheets may not be the secondary structure most characteristic of native phosvitin (physiological pH), even though beta-sheet is the predominant conformation for phosvitin in acidic solutions (pH 1.5) and in the lyophilized solid. Phosvitin appears to be the first protein for which the major component to the Raman amide I band is centered near 1685 cm-1, which is 10-40 cm-1 higher than proteins heretofore examined in aqueous solution by Raman spectroscopy. |
