Immunoreactivity and conformation of the P-P-G-M-R-P-P repetitive epitope of the Sm autoantigen.
Anti-Sm antibodies are usually considered highly specific for systemic lupus erythematosus (SLE), while anti-U1 RNP antibodies are found in high titers in patients with mixed connective tissue disease (MCTD). The sequence P1-P-G-M-R-P-P7, present in three copies in the Sm (U1-U6 RNA-protein complex) autoantigen, is an important functional domain of the antigenic determinants. The immunoreactivity of this proline-rich repetitive epitope was investigated by testing sera with various autoantibody specificities for reactivity against this epitope, as well as its conformational properties by means of 1D and 2D 1HNMR spectroscopy. It was found that the P-P-G-M-R-P-P epitope is recognized mainly by anti-U1RNP and/or anti-Sm positive sera, but also by anti-Ro(SSA) (hY1RNA-protein complex) and anti-La(SSB) (hY1RNA-protein complex) positive sera, although these sera are negative for anti-U1RNP and anti-Sm. Conformational analysis of the proline-rich epitope in DMSO-d6 solution obtained from lyophilized aqueous solution at pH 5 showed the presence of at least three conformers. The main conformer A (62%) is stabilized by an ionic interaction between the guanidinium and the C-terminal carboxylate groups, and the Pro6-Pro7 peptide bond adopts the cis form. A type II beta-turn is also present in the N-terminal sequence (Pro1-Pro-Gly-Met4-) of this conformer. Conformer B (21%) is also stabilized by a similar ionic interaction, as in conformer A, while the NMR data indicate the absence of a folded structure in the N-terminal tetrapeptide of this conformer. Conformer C (17%) adopts a completely extended structure. The multiple conformers of the P-P-G-M-R-P-P may offer some explanation for the reactivity of sera with various autoantibody specificities against this epitope.
Tsikaris
V
V
Department of Chemistry, University of Ioannina, Greece.
Vlachoyiannopoulos
P G
PG
Panou-Pomonis
E
E
Marraud
M
M
Sakarellos
C
C
Moutsopoulos
H M
HM
Sakarellos-Daitsiotis
M
M
D000595:Amino Acid Sequence; D001323:Autoantibodies / Q000097:blood; D001324:Autoantigens / Q000737:chemistry* / Q000276:immunology; D001327:Autoimmune Diseases / Q000097:blood / Q000276:immunology*; D001667:Binding, Competitive; D002851:Chromatography, High Pressure Liquid; D003240:Connective Tissue Diseases / Q000097:blood / Q000276:immunology*; D004121:Dimethyl Sulfoxide / Q000737:chemistry; D004797:Enzyme-Linked Immunosorbent Assay; D000939:Epitopes / Q000032:analysis / Q000737:chemistry* / Q000276:immunology; D006801:Humans; D006863:Hydrogen-Ion Concentration; D008180:Lupus Erythematosus, Systemic / Q000097:blood / Q000276:immunology; D009682:Magnetic Resonance Spectroscopy; D008947:Mixed Connective Tissue Disease / Q000097:blood / Q000276:immunology; D009842:Oligopeptides / Q000032:analysis / Q000737:chemistry* / Q000276:immunology; D011487:Protein Conformation; D011522:Protons
D001323:Autoantibodies; D001324:Autoantigens; D000939:Epitopes; D009842:Oligopeptides; D011522:Protons; D004121:Dimethyl Sulfoxide