Identification and chemical synthesis of a ribosomal protein antigenic determinant in systemic lupus erythematosus.

Abstract

The characteristics of eukaryotic ribosomal proteins PO, P1, and P2 (P proteins) and their antigenic determinants were studied using the sera of patients with systemic lupus erythematosus (SLE). PO, P1, and P2 were isolated as a macromolecular complex by preparative isoelectric focusing and anion-exchange chromatography in the-presence of 6 M urea. The apparent molecular size of the complex was 140 kDa as determined by gel fitration on a Sephadex G-200 column. PO may, therefore, be the eukaryotic equivalent ofEscherichia coli ribosomal protein L10. In addition, all three P proteins were detected in the postribosomal supernatant of HeLa cells, and PO and P1 were found to be more acidic than their ribosome-bound counterparts. Partial proteolysis experiments revealed that SLE anti-P sera recognized one or both ends of the P2 equivalent protein from Artemia saUna (eL12). Sixteen SLE sera containing antibodies to PO, P1, and P2 reacted with a carboxyl-terminal peptide 22 amino acids in length of eL12 and not with an amino-terminal peptide of 20 anmno acids. Even though the carboxyl-terminal peptide completely inhibited the ability of the antiserum to react with all three proteins on an immunological blot, the same peptide produced only small decreases in binding of the SLE antibody to the native, nondenatured P proteins. These findings indicate that SLE anti-P antibodies react with a single sequential (linear) antigenic determinant on all three P proteins, but that additional antibodies recognize a conformational determinant(s).Systemic lupus erythematosus (SLE) is an autoimmune disease generally characterized by serum antibodies directed against nuclear proteins and nucleic acids (1). That some patients' sera contain antibodies against ribosomal constituents (2, 3) has long been known, but the identity of these ribosomal antigens has only recently been determined (4). SLE anti-ribosome antibodies show almost exclusive reactivity against three 60S ribosomal subunit phosphoproteins called PO, P1, and P2 (4, 5). These same three proteins are also recognized by a mouse monoclonal antibody raised against chicken ribosomes (6). These "P" proteins have molecular sizes of -38, 19, and 17 kDa, respectively. P1 and P2 are believed to be the eukaryotic equivalent of the Escherichia coli ribosomal protein L12 and have been shown to contain sequences that are highly conserved among eukaryotes (6). Thus, P2 (7) from rat liver shows a high degree of amino acid sequence homology with Artemia salina ribosomal protein eL12 (8) and yeast ribosomal protein YPA1 (9). P1 and P2 also appear to be the functional counterparts of Artemia ribosomal proteins eL12' and eL12 and yeast ribosomal proteins YPA1/YPA2 (10-12). In order to further evaluate the remarkable specificity of the SLE anti-PO, -P1, and -P2 (anti-P) antibodies, we have mapped the antigenic determinant on the P proteins. This determinant is present on all three proteins and is contained within a common sequence of 22 amino acids at the carboxyl terminus of the Artemia r...

Anti–ribosomal P protein antibody in human systemic lupus erythematosus up‐regulates the expression of proinflammatory cytokines by human peripheral blood monocytes

“…Autoantibodies to ribosomal P proteins (anti-P antibodies) are detected in 12-16% of patients with SLE (1). Anti-P antibodies are directed to 3 phosphoproteins (P0, P1, and P2), which are located on the larger 60S subunit of eukaryotic ribosomes and have molecular weights of 38 kd, 19 kd, and 17 kd, respectively (1). Ribosomal P proteins share a common linear determinant that is present in the carboxyl-terminal 22-amino acid sequence (1).…”

“…Anti-P antibodies are directed to 3 phosphoproteins (P0, P1, and P2), which are located on the larger 60S subunit of eukaryotic ribosomes and have molecular weights of 38 kd, 19 kd, and 17 kd, respectively (1). Ribosomal P proteins share a common linear determinant that is present in the carboxyl-terminal 22-amino acid sequence (1). Previous and recent studies have disclosed the association of anti-P antibodies with neuropsychiatric disease in SLE (2,3).…”

Binding of Mammalian Ribosomal Protein Complex P0·P1·P2 and Protein L12 to the GTPase-associated Domain of 28 S Ribosomal RNA and Effect on the Accessibility to Anti-28 S RNA Autoantibody

“…To know whether this consisted of the three P proteins, P0, P1, and P2, protein components were isolated from this band, subjected to SDS-gel electrophoresis, and then followed by immunoblotting with anti-P antibody. This antibody recognizes a common carboxyl-terminal epitope of the three P proteins (Elkon et al, 1986). As shown in Fig.…”

Association of IgG anti‐NR2 glutamate receptor antibodies in cerebrospinal fluid with neuropsychiatric systemic lupus erythematosus

“…The synthetic DWEYSVWLSN peptide, of which DWEYS corresponds to human NMDA receptors NR2a and NR2b (amino residues 283-287), was synthesized using a solid-phase method (Peptide Institute, Minoh, Osaka, Japan) as previously described (8). The purity of the peptide was confirmed as 98.1%, using analytic high-performance liquid chromatography, amino acid analysis, and microsequencing.…”