Identification of the epitope of a monoclonal antibody to DJ-1.
Mutations in DJ-1 can cause early onset parkinsonism. Various antibodies have been generated to detect this protein, one of which is a commonly used monoclonal antibody (clone 3E8). Since results of in situ examinations of DJ-1 expression with this antibody have differed from analyses with species-specific antibodies (e.g. rat), it would be useful to know the epitope for this antibody. Using GFP-tagged deletion constructs of human DJ-1, we have localized the epitope region for this antibody to within residues 56-78 of human DJ-1. Mapping this region to the published three-dimensional structure of DJ-1 indicates that this is a solvent-accessible surface epitope. Immunonegativity of E64D mutant DJ-1 with the monoclonal antibody suggests that glutamate 64 of human DJ-1 contributes to the epitope recognized by this antibody. Moreover, the loss of immunoreactivity due to such a small substitution demonstrates the remarkable sensitivity of the monoclonal antibody 3E8 to DJ-1.
Miller
David W
DW
Laboratory of Neurogenetics, National Institute on Aging, Bldg 35, Rm 1A-1002, 35 Convent Drive, Bethesda, MD 20892, USA. millerda@mail.nih.gov
Wilson
Carmen R
CR
Kaleem
Mona A
MA
Blackinton
Jeff
J
Cookson
Mark R
MR
D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal / Q000737:chemistry*; D000918:Antibody Specificity; D000937:Antigen-Antibody Reactions; D015153:Blotting, Western / Q000379:methods; D002460:Cell Line; D000939:Epitopes / Q000276:immunology; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins; D051379:Mice; D016297:Mutagenesis, Site-Directed / Q000276:immunology; D015513:Oncogene Proteins / Q000276:immunology*; D000071617:Protein Deglycase DJ-1; D017433:Protein Structure, Secondary; D051381:Rats; D014162:Transfection / Q000379:methods
D000911:Antibodies, Monoclonal; D000939:Epitopes; D047908:Intracellular Signaling Peptides and Proteins; D015513:Oncogene Proteins; C105131:PARK7 protein, human; D000071617:Protein Deglycase DJ-1