Synthetic peptides from region 65-84 of bovine myelin basic protein: radioimmunoassays and equilibrium competitive inhibition studies with antibodies prepared against myelin basic protein
Day, E.D.; Hashim, G.A.; Varitek, V.A.; Lazarus, K.J.; Paterson, P.Y.
Neurochemical Research 6(8): 913-929
Synthetic peptides with various and overlapping sequences represented by the residue region 65-84 of bovine myelin basic protein (MBP-bov) were tested in Na2SO4 radioimmunoassays [RIA] for their reactivity with 15 rabbit antisera against MBP from 6 different animal speceis and 9 pools of syngeneic Lewis rat anti-MBP antisera. Three of the peptides were labeled with 125I and studied by direct binding reactions: the prototype peptide S82 sequence TTHYGSLPQKAQGHRPQDEG, corresponding to residues 65-84 of bovine MBP except for the C-terminal glycine, which is encephalitogenic in rabbits; S81, which lacks the first 3 residues of S82 and is nonencephalitogenic in rabbits; and S79, which represents the C-terminal half of S82 and which, because of its C-terminal glycine instead of asparagine, is nonencephalitogenic in Lewis rats. Of the rabbit anti-MBP antisera 14 were reactive with [125I]S82 (2 borderline) including 2 of 3 antisera against human MBP, 1 against monkey MBP, and 1 against chicken MBP. The cross-reactions with [125I]S81 were somewhat fewer and less intense. There were no cross-reactions with [125I]S79. None of 9 different pools of syngeneic rat MBP antisera cross-reacted with any of the 3 labeled peptides. With the use of a rabbit anti-MBP-rat antiserum that cross-reacted strongly with [125I]S82, 15 additional peptides with overlapping sequences within the residue region 65-84, and 5 MBP preparations from 4 different species, were tested by equilibrium and nonequilibrium competitive inhibition RIA. Unlabeled S82 and MBP-bov were completely competitive with [125I]S82 in the equilibrium assays; S81 and 3 other peptides had low degrees of cross-reativity; but none of the remaining 8 unlabeled peptides or unlabeled MBP preparations of guinea pig, rat or mouse origin gave any evidence of competitive activity. Nonequilibrium competitive inhibition RIA did reveal cross-reactivities among several of the peptides and guinea pig and rat MBP. The N-terminal half of S82, particularly residues 68-74 (YGSLPQK), must apparently contain an immunodeterminant of amino acid residues which identifies with the corresponding and exposed sequence in intact MBP-bov.