Abstract
The Ku (p70/p80) autoantigen consists of two phosphoproteins of molecular mass approximately 70,000 and 80,000 forming a macromolecular complex that binds DNA. Autoantibodies from a patient with systemic lupus erythematosus were used to isolate cDNA clones encoding the human approximately 70-kDa Ku antigen (p70) from a lambda gt11 expression library. The deduced amino acid sequence of p70 consisted of 609 amino acid residues and was confirmed by partial amino acid sequencing. The protein contains two acidic domains of 61 residues (31% Glu + Asp) and 19 residues (53% Glu + Asp) that are similar in size and charge to those found in a number of proteins involved in transcriptional activation. The 61-residue acidic region is rich in serine, raising the possibility that its charge might be modulated by phosphorylation. The predicted amino acid sequence also contains two regions with periodic repeats of either leucine alone, or leucine alternating with serine every seventh position. The latter repeat displays sequence and secondary structural similarities with the "leucine zipper" regions of the c-myc and v-myc oncogene products. The p70 antigen does not appear to have extensive sequence homology with the 80-kDa Ku autoantigen based on analysis of RNA blots and immunological criteria. A major antigenic determinant or determinants recognized by human autoantibodies is located near a leucine repeat on the carboxyl-terminal 190 amino acid residues of p70.
Highlights
The Ku (p70/p80) autoantigenconsists of two phos- chromosomes in early prophase [2]
We have used autoantibodies from the of p70 consisted of 609 amino acid residues and was serum of an individual with systemic lupus erythematosus (SLE) to isolate cDNA clones
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Summary
Antibodies specific for the fusion proteins were purified by elution from the nitrocellulose blots [18] and used to probe immunoblots of K562 nuclear extract [2] followed by detection with 1261protein A as described above. Sequence of p70 wascompared to sequences in the National Biomed- Nucleicacid hybridization screening yielded additional ical Research Foundation Protein Identification Resource (PIR) us- Xgtll clones hybridizing with both the clone 70.77 insert and ing the algorithms of Lipman and Pearson [24,25]. The affinity-purified anti-70.34 and anti70.77 antibodies bound to p70 on immunoblots of total nuclear proteins, while autoantibodies in the original CK serum bound to both p70 and p80 (Fig. 3A).
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