Localization of a conformational epitope common to non-native and fibrillar immunoglobulin light chains.
Amyloid fibrils and partially unfolded intermediates may be distinguished serologically from native amyloidogenic precursor proteins or peptides. In this regard, we had previously reported that the IgG1 mAb 11-1F4, generated by immunizing mice with a thermally denatured variable region fragment of the human Igkappa4 Bence Jones protein Len, reacted specifically with light chain (LC) fibrils, irrespective of kappa or lambda isotype but, notably, did not with native molecules (Hrncic, R. et al. (2000) Am. J. Pathol. 157, 1239-1246). To elucidate the molecular basis of this specificity, we have used a europium-linked fluorescent immunoassay, where it was demonstrated through epitope mapping that mAb 11-1F4 recognizes a conformational determinant contained within the first (N-terminal) 18 amino acids of misfolded LCs. The nature of this epitope was evidenced in competition studies where the peptide Len (1-18), but not the intact protein or other LCs, inhibited the binding of the antibody to fibrils. This unique reactivity was dependent on the structural integrity of this portion of the molecule, particularly the presence of a highly conserved prolyl residue at position 8. On the basis of our experimental data, we posit that the mAb 11-1F4 binding site found on partially denatured and fibrillar LCs involves an inducible N-terminal main chain reversal that results in the formation of a proline anchored beta-turn. Our delineation of this LC fibril-associated epitope provides the rationale for the design of novel amyloid-reactive antibodies with diagnostic and therapeutic potential for patients with LC-associated and other forms of amyloidosis.
O'Nuallain
Brian
B
Human Immunology and Cancer Program, Department of Medicine, University of Tennessee Graduate School of Medicine, Knoxville, Tennessee 37920, USA.
Allen
Amy
A
Kennel
Stephen J
SJ
Weiss
Deborah T
DT
Solomon
Alan
A
Wall
Jonathan S
JS
D000682:Amyloid / Q000737:chemistry / Q000276:immunology* / Q000302:isolation & purification; D000686:Amyloidosis / Q000276:immunology*; D000911:Antibodies, Monoclonal / Q000276:immunology; D000918:Antibody Specificity; D018604:Epitope Mapping; D000939:Epitopes; D006801:Humans; D007147:Immunoglobulin Light Chains / Q000737:chemistry* / Q000276:immunology*; D011487:Protein Conformation; D011489:Protein Denaturation / Q000276:immunology
D000682:Amyloid; D000911:Antibodies, Monoclonal; D000939:Epitopes; D007147:Immunoglobulin Light Chains