Antigenic epitopes of the photoreceptor synaptic ribbon.
The purpose of this study was twofold: 1) to purify and identify a protein containing an epitope recognized by an anti-synaptic ribbon antibody B16 and 2) to identify and sequence the epitope. B16 recognizes several unrelated proteins in retina immunoblots. Purification and microsequencing of the strongest band (88 kDa) demonstrate 94% identity to aconitase over 111 amino acids. Polyclonal antibodies against aconitase recognize aconitase on Western blots, but not synaptic ribbons in sections. We conclude that although aconitase contains the epitope, aconitase is not the synaptic ribbon protein. The B16 epitope was identified to be 542DTYQHPPKDS551. A synthetic peptide to this sequence absorbs B16 activity in both Western blots and immunohistochemistry studies, whereas partial peptides fail to absorb activity. Additional antibodies against this peptide label synaptic ribbons. When mouse retina were double labeled with B16 and anti-alpha-actinin, B16 was found to label synaptic ribbons in the outer plexiform layer that partially enclosed the alpha-actinin label. We have determined the amino acid sequence of the B16 epitope and found that the B16 labeling colocalizes with alpha-actinin at the photoreceptor synapse.
Nguyen
T H
TH
Child Study Center, Yale University School of Medicine, New Haven, Connecticut 06520, USA.
Balkema
G W
GW
D000154:Aconitate Hydratase / Q000276:immunology; D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology*; D003429:Cross Reactions; D000939:Epitopes / Q000276:immunology / Q000302:isolation & purification*; D005136:Eye Proteins / Q000276:immunology*; D006801:Humans; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D010446:Peptide Fragments / Q000737:chemistry / Q000276:immunology; D020419:Photoreceptor Cells, Vertebrate / Q000276:immunology*; D011817:Rabbits; D011993:Recombinant Fusion Proteins / Q000276:immunology; D013569:Synapses / Q000276:immunology*
D000911:Antibodies, Monoclonal; D000939:Epitopes; D005136:Eye Proteins; D010446:Peptide Fragments; D011993:Recombinant Fusion Proteins; D000154:Aconitate Hydratase