Deposition of amyloid fibrils consisting of amyloid beta (A beta) protein as senile plaques in the brain is a pathological hallmark of Alzheimer's disease. However, a growing body of evidence shows that soluble A beta oligomers correlate better with dementia than fibrils, suggesting that A beta oligomers may be the primary toxic species. The structure and oligomerization mechanism of these A beta oligomers are crucial for developing effective therapeutics. Here we investigated the oligomerization of A beta 42 in the context of a fusion protein containing GroES and ubiquitin fused to the N-terminus of A beta sequence. The presence of fusion protein partners, in combination with a denaturing buffer containing 8 M urea at pH 10, is unfavorable for A beta 42 aggregation, thus allowing only the most stable structures to be observed. Transmission electron microscopy showed that A beta 42 fusion protein formed globular oligomers, which bound weakly to thioflavin T and Congo red. SDS-PAGE shows that A beta 42 fusion protein formed SDS-resistant hexamers and tetramers. In contrast, A beta 40 fusion protein remained as monomers on SDS gel, suggesting that the oligomerization of A beta 42 fusion protein is not due to the fusion protein partners. Cysteine scanning mutagenesis at 22 residue positions further revealed that single cysteine substitutions of the C-terminal hydrophobic residues (I31, I32, L34, V39, V40, and I41) led to disruption of hexamer and tetramer formation, suggesting that hydrophobic interactions between these residues are most critical for A beta 42 oligomerization. (C) 2011 Elsevier Inc. All rights reserved.