The monomer to oligomer transition initiates the aggregation and pathogenic transformation of Alzheimer amyloid-␤ (A␤) peptide. However, the monomeric state of this aggregationprone peptide has remained beyond the reach of most experimental techniques, and a quantitative understanding of this transition is yet to emerge. Here, we employ single-molecule level fluorescence tools to characterize the monomeric state and the monomer-oligomer transition at physiological concentrations in buffers mimicking the cerebrospinal fluid (CSF). Our measurements show that the monomer has a hydrodynamic radius of 0.9 ؎ 0.1 nm, which confirms the prediction made by some of the in silico studies. Surprisingly, at equilibrium, both A␤ 40 and A␤ 42 remain predominantly monomeric up to 3 M, above which it forms large aggregates. This concentration is much higher than the estimated concentrations in the CSF of either normal or diseased brains. If A␤ oligomers are present in the CSF and are the key agents in Alzheimer pathology, as is generally believed, then these must be released in the CSF as preformed entities. Although the oligomers are thermodynamically unstable, we find that a large kinetic barrier, which is mostly entropic in origin, strongly impedes their dissociation. Thermodynamic principles therefore allow the development of a pharmacological agent that can catalytically convert metastable oligomers into nontoxic monomers.Alzheimer disease (AD) 2 is a degenerative brain disorder that is associated with the presence of extracellular aggregates of amyloid-␤ (A␤) (1), which is an ϳ4.5-kDa peptide containing 39 -42 residues. Recent studies indicate that small soluble oligomers are key to A␤ toxicity (2-4). In the AD brain, both A␤ monomers and dimers have been isolated, and the dimers have been shown to impair synaptic plasticity in mouse hippocampal slices (5). In contrast, A␤ monomers have been shown to be devoid of neurotoxicity (5) and have in fact been suggested to be neuroprotective (6, 7). The monomer to oligomer transition is therefore not only the obligatory first event of aggregation, it is also the key event determining the transformation of a benign protein to a neurotoxic one.We address this transition from a thermodynamic viewpoint: an aggregation-capable molecule should have a defined equilibrium between monomers and dimers (or oligomers), such that it is primarily monomeric below a certain concentration. Any oligomer-enriched solution prepared below such a concentration must be thermodynamically unstable and must dissociate to monomers at a given rate. To understand AD in terms of A␤ aggregation, we need to understand how this concentration compares with the in vivo concentrations of A␤ (which is estimated to be Ͻ Ͻ1 M) (8 -11) and what the kinetics of A␤ oligomer dissociation is.However, experiments probing the monomer to oligomer transition have been difficult to perform due to the low concentration at which this transition most likely occurs, and they have yielded rather confusing results. Some studies have ...

The transcriptional regulator p53 has an essential role in tumor suppression. Almost 50% of human cancers are associated with the loss of p53 functions, where p53 often accumulates in the nucleus as well as in cytoplasm. Although it has been previously suggested that amyloid formation could be a cause of p53 loss-of-function in subset of tumors, the characterization of these amyloids and its structure-function relationship is not yet established. In the current study, we provide several evidences for the presence of p53 amyloid formation (in human and animal cancer tissues); along with its isolation from human cancer tissues and the biophysical characterization of these tissue-derived fibrils. Using amyloid seed of p53 fragment (P8, p53(250-257)), we show that p53 amyloid formation in cells not only leads to its functional inactivation but also transforms it into an oncoprotein. The in vitro studies further show that cancer-associated mutation destabilizes the fold of p53 core domain and also accelerates the aggregation and amyloid formation by this protein. Furthermore, we also show evidence of prion-like cell-to-cell transmission of different p53 amyloid species including full-length p53, which is induced by internalized P8 fibrils. The present study suggests that p53 amyloid formation could be one of the possible cause of p53 loss of function and therefore, inhibiting p53 amyloidogenesis could restore p53 tumor suppressor functions.

Background: Aggregation of ␣-Syn is associated with PD pathogenesis. Results: Despite being natively unfolded, a site-specific structure exists in ␣-Syn that is significantly altered by familial PD-associated E46K, A53T, and A30P mutations. Conclusion: Altered site-specific structure of the PD-associated mutants may attribute to their different aggregation propensity. Significance: This study contributes to understanding the relationship between structure and aggregation of ␣-Syn.

Background: Peptide/protein hormones are stored as amyloids within endocrine secretory granules. Results: Disulfide bond cleavage enhances conformational dynamics and aggregation kinetics in somatostatin-14, resulting in amyloid fibrils with increased resistance to denaturing conditions and decreased reversibility. Conclusion: Disulfide bond could be a key modulating factor in somatostatin-14 amyloid formation associated with secretory granule biogenesis. Significance: Defective disulfide bonding might cause dysregulation of hormone storage/secretion.

The serotonin(1A) receptor is a representative member of the G-protein coupled receptor (GPCR) superfamily and serves as an important target in the development of therapeutic agents for neuropsychiatric disorders. Oligomerization of GPCRs is an important contemporary issue since it is believed to be a crucial determinant for cellular signaling. In this work, we monitored the oligomerization status of the serotonin(1A) receptor tagged to enhanced yellow fluorescent protein (5-HT(1A)R-EYFP) in live cells utilizing time-resolved fluorescence anisotropy decay. We interpret the unresolved fast component of the observed anisotropy decay as fluorescence resonance energy transfer (FRET) between 5-HT(1A)R-EYFP molecules (homo-FRET). Homo-FRET enjoys certain advantages over hetero-FRET in the analysis of receptor oligomerization. Our results reveal the presence of constitutive oligomers of the serotonin(1A) receptor in live cells. We further show that the oligomerization status of the receptor is independent of ligand stimulation and sphingolipid depletion. Interestingly, acute (but not chronic) cholesterol depletion appears to enhance the oligomerization process. Importantly, our results are independent of receptor expression level, thereby ruling out complications arising due to high expression. These results have potential implications in future therapeutic strategies in pathophysiological conditions in which serotonin(1A) receptors are implicated.