Inherent Anti-amyloidogenic Activity of Human Immunoglobulin γ Heavy Chains

Brian O'Nuallain,Sharad P Adekar,Alan Solomon,Michael J Rowan,Sally Macy,Scott K Dessain,Igor Klyubin

doi:10.1074/jbc.m109.044321

Abstract

We have previously shown that a subpopulation of naturally occurring human IgGs were cross-reactive against conformational epitopes on pathologic aggregates of Abeta, a peptide that forms amyloid fibrils in the brains of patients with Alzheimer disease, inhibited amyloid fibril growth, and dissociated amyloid in vivo. Here, we describe similar anti-amyloidogenic activity that is a general property of free human Ig gamma heavy chains. A gamma(1) heavy chain, F1, had nanomolar binding to an amyloid fibril-related conformational epitope on synthetic oligomers and fibrils as well as on amyloid-laden tissue sections. F1 did not bind to native Abeta monomers, further indicating the conformational nature of its binding site. The inherent anti-amyloidogenic activity of Ig gamma heavy chains was demonstrated by nanomolar amyloid fibril and oligomer binding by polyclonal and monoclonal human heavy chains that were isolated from inert or weakly reactive antibodies. Most importantly, the F1 heavy chain prevented in vitro fibril growth and reduced in vivo soluble Abeta oligomer-induced impairment of rodent hippocampal long term potentiation, a cellular mechanism of learning and memory. These findings demonstrate that free human Ig gamma heavy chains comprise a novel class of molecules for developing potential therapeutics for Alzheimer disease and other amyloid disorders. Moreover, establishing the molecular basis for heavy chain-amyloidogenic conformer interactions should advance understanding on the types of interactions that these pathologic assemblies have with biological molecules.

Highlights

Tein misfolding diseases that are termed the amyloidoses [1, 2]
Among these molecules are a subpopulation of A␤-reactive polyclonal IgGs in intravenous immune globulin, derived from pools of plasma from presumably healthy donors, that we have shown to cross-react with amyloid fibrils and oligomers [17, 18]
Having established that heavy chain (HC) F1 bound to a conformational epitope present on synthetic amyloid fibrils and oligomers, we investigated the ability of the protein to bind to patientderived amyloid fibrils and to stain amyloid-laden patient tissue sections

Summary

EXPERIMENTAL PROCEDURES

Proteins, and Antibodies—Human A␤40, A␤42, and human islet amyloid polypeptide (IAPP) were purchased from Quality Controlled Biochemicals (Hopkinton, MA). Antibodies or HCs were serially diluted in assay buffer (1% BSA in PBSA) and tested (100 ␮l/well) in activated, high binding microtiter plate wells (COSTAR, Corning, NY) that were coated with 400 ng of target protein and blocked with 1% BSA (Sigma-Aldrich) in PBSA. For measurements of fibril extension, HCs F1 and 13A or IgG 13A were serially diluted (10 nM-1.0 ␮M) in PBSA in high binding microtiter plate wells (COSTAR; Corning) that were coated with 400 ng of sonicated A␤ fibrils. Two independent hybridomas, F1 and F2, were isolated that bound to LC fibrils with molar antibody concentrations that obtained half-maximum binding (EC50) values of ϳ20 nM (Fig. 1 and Table 1). F1 bound to CAPS as strongly as to amyloid fibrils, with an EC50 value of ϳ20 nM, and had weak (␮M) interactions with plate-

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Findings

DISCUSSION