Identification of an Inhibitory Element within the Human 68-kDa (U1) Ribonucleoprotein Antigen
| Autor: | Nicholas Hunt, Michael Walter, Heike Berg, Wolfgang Northemann, Steve Fenning, Gisela Stahnke |
|---|---|
| Rok vydání: | 1995 |
| Předmět: |
DNA
Complementary Molecular Sequence Data Gene Expression Biology Autoantigens Ribonucleoprotein U1 Small Nuclear law.invention Antigen law Complementary DNA Gene expression Escherichia coli Humans Amino Acid Sequence Northern blot Cloning Molecular Peptide sequence Autoantibodies Mixed Connective Tissue Disease Ribonucleoprotein Immunoassay Base Sequence Chromosome Mapping Molecular biology Recombinant Proteins Molecular Weight Recombinant DNA Small nuclear ribonucleoprotein Plasmids Biotechnology |
| Zdroj: | Protein Expression and Purification. 6:748-756 |
| ISSN: | 1046-5928 |
| DOI: | 10.1006/prep.1995.0005 |
| Popis: | Various nuclear proteins are the major targets of autoimmune responses in various rheumatic disorders. In particular, autoantibodies directed against a 68-kDa protein associated with the (U1) RNA-containing small nuclear ribonucleoprotein complexes typically occur in sera of patients with mixed connective tissue disease and related rheumatic disorders, such as systemic lupus erythematosus, and therefore are very useful as a serological marker. For establishing powerful immunoassays, it was necessary to generate recombinant human P68 antigen as the antigenic target. In this study we demonstrated that the cDNA coding for the full-length human P68 antigen could not be expressed by a traditional bacterial vector system due to a putative inhibitory sequence designated as inhibitory sequence X which is located between the autoreactive domains C′ and D′ of the human P68 antigen. The construction of corresponding hybrid plasmids carrying two functional and independent gene blocks indicated the trans -active function of the inhibitory sequence X, which could be localized by expression studies of various deletion constructs. Comparable Northern blot analysis clearly demonstrated that the inhibitory sequence X could act on the translation of the P68 mRNA. After excision of the inhibitory sequence X a dramatic increase in the production of recombinant human P68 antigen was observed. |
| Databáze: | OpenAIRE |
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