Molecular Characterization of the Ro/SS-A Autoantigens

AbstractMolecular techniques have recently revealed that there are several immunologically distinct Ro/SS-A antigens. Three genes encoding putative Ro/SS-A protein antigens with calculated masses of 46, 52, and 60 kD have been isolated. The encoded amino acid sequence of each is quite dissimilar. The 46-kD antigen is calreticulin (CR), a highly conserved calcium-binding protein that resides predominately in the endoplasmic reticulum where it may be involved in protein assembly. Although CR has recently been confirmed to be a new human rheumatic disease-associated autoantigen, its relationship to the other components of the Ro/SS-A ribonucleoprotein has become somewhat controversial owing predominately to the fact that recombinant forms of calreticulin have not displayed the same pattern of autoantibody reactivity possesse by the native form of this protein.The 52-kD antigen most likely resides in the nucleus and may be involved in the regulation of gene expression. The cellular location and function of the 60-kD antigen is uncertain but studies indicate that it is a RNA-binding protein.The 46- and 60-kD antigens share homology with foreign polypeptides, suggesting that an immune response initially directed against a foreign protein may give rise to the autoimmune response directed at cross-reacting self proteins

Primary structure of the N-linked carbohydrate chains of calreticulin from spinach leaves

Calreticulin is a multifunctional Ca2+-binding protein of the endoplasmic reticulum of most eukaryotic cells. The 56 kDa Calreticulin glycoprotein isolated from spinach (Spinacia oleracea L.) leaves was N-deglycosylated by PNGase-F digestion. The carbohydrate moiety was isolated by gel permeation chromatography and purified by high-pH anion-exchange chromatography. The fractions were investigated by 500 MHz H-1-NMR spectroscopy, in combination with monosaccharide analysis and fast-atom bombardment-mass spectrometry. The following carbohydrate structure could be established as the major component (Man(8)GlcNAc(2)): [GRAPHICS] Heterogeneity was demonstrated by the presence of two minor components being Man(7)GlcNAc(2) lacking a terminal residue (D-1 or D-3), compared to the major component. A. cross-reactivity with an antibody against the endoplasmic reticulum retention signal HDEL was also found