Ultraviolet cross-linking of helical oligonucleotides to two monoclonal MRL-1pr/1pr anti-DNA autoantibodies. Variations in H and L chain binding to DNA
- Tufts Univ., Boston, MA (USA)
Experiments were performed to determine whether both H and L chains of different anti-native DNA autoantibodies are uniformly involved in binding to DNA. Two purified monoclonal mouse (MRL-1pr/1pr) IgG autoantibodies, H241 and 2C10, were tested. They both bound synthetic helical oligonucleotides of 10 to 20 base pairs in a gel electrophoresis retardation assay but differed in their preferences for given base sequences. Exposure of antibody-radiolabeled oligonucleotide mixtures to UV light (254 nm) for 10 min led to specific covalent cross-linking of oligonucleotide to both the H and the L chains of H241 but only to the H chain of 2C10. Single labeling events were detected without higher aggregation. The oligonucleotides were not cross-linked to unrelated IgG, even after 2 h of irradiation. Cross-linked (radioactively labeled) H and L chains of H241 and 2C10 were isolated from denaturing electrophoresis gels and digested with lysyl endopeptidase and/or staphylococcal V8 protease. H241 and 2C10 H chains each yielded a major labeled peptide fragment, but the peptides from the two antibodies were different. These experiments measured only some of the antibody-DNA interactions, probably with bases in the major groove of the DNA. They indicated that two MRL-1pr/1pr IgG anti-native DNA antibodies differ in their H and L chain contacts with DNA and provide an approach to identifying affinity-labeled binding sites in the antibodies.
- OSTI ID:
- 6168351
- Journal Information:
- Journal of Immunology; (USA), Vol. 145:10; ISSN 0022-1767
- Country of Publication:
- United States
- Language:
- English
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Related Subjects
OLIGONUCLEOTIDES
CHEMICAL RADIATION EFFECTS
CROSS-LINKING
DNA
ELECTROPHORESIS
MICE
MONOCLONAL ANTIBODIES
ULTRAVIOLET RADIATION
ANIMALS
ANTIBODIES
CHEMICAL REACTIONS
CHEMISTRY
ELECTROMAGNETIC RADIATION
MAMMALS
NUCLEIC ACIDS
ORGANIC COMPOUNDS
POLYMERIZATION
RADIATION CHEMISTRY
RADIATION EFFECTS
RADIATIONS
RODENTS
VERTEBRATES
560120* - Radiation Effects on Biochemicals
Cells
& Tissue Culture