Single-Molecule Atomic Force Microscopy Force Spectroscopy Study of Aβ-40 Interactions

Kim, Bo Hyun; Palermo, Nicholas Y.; Lovas, Sándor; Zaikova, Tatiana; Keana, John F. W.; Lyubchenko, Yuri L.

doi:10.1021/bi200147a

Cited by 81 publications

(126 citation statements)

References 48 publications

(155 reference statements)

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“…Furthermore, such conformations are characteristic of the dimers. This finding is in agreement with the dynamic force spectroscopy study (DFS) which predicted the presence of stable dimers in solution with a life-time of 0.1 s at pH 7.0 [6]. The DFS analysis of A(1-40) dimers also showed that various pathways of dimer dissociation occur indicating that the dimers are built with different conformations.…”

supporting

confidence: 89%

Misfolding and Oligomerization of Amyloid β(1-40)

Lovas¹,

Watts²

2015

Peptides 2015, Proceedings of the 24th American Peptide Symposium

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IntroductionProtein misfolding and self-assembly have been identified as important factors in the development of several neurodegenerative diseases. The hallmark of many of these diseases is the deposition of amyloid fibrils and plaques within the extracellular matrix with an organized core structure consisting of -sheets running perpendicular to the fibril axis. The histologic sign of Alzheimer's disease is the presence of senile plaques formed from Amyloid (A) and neurofibrillary tangles formed from hyperphosphorylated tau protein. Despite a more thorough understanding of fibril structure, A belongs to an inherently disordered class of polypeptides making its structural determination in both monomeric and oligomeric forms difficult. Previous studies have been dependent on maintaining peptide solubility through the use of organic solvents or detergents which would potentially have significant effects on the overall solution structure of the peptide. Recently, a solution NMR structure of A(1-40) in 50 mM NaCl solution has been determined (PDB id.: 2LFM) [1]. The molecular mechanism of dimerization, subsequent oligomerization and the conformation of the multimeric peptide chains are still not known. Therefore, herein we describe the structural properties of the dimer of A(1-40) using 300 ns replica exchange molecular dynamics (REMD) simulation in TIP3P water as solvent using the CHARMM36 force field as implemented in the GROMACS version 4.6.4 package [2]. The dimer structure was created by randomly docking the NMR-derived solution structure with itself and relaxed with a 1.4 s MD simulation. The REMD simulations were performed with 70 replicas of the initial structure. The exchange probability was 0.2. The exchange temperatures were obtained from a web-based temperature predictor [3] with the lower and upper limits of 290 K and 470 K, respectively. Results and DiscussionThe backbone conformational entropy for the dimer system was calculated for all replica exchange temperatures. Two important aspects of the dimer system are evident from examination of the evolution of entropy during simulations. First, a relatively long, approximately 100000 ps, period of simulation time is required for the system to reach equilibrium. Second, the structures sampled at 310K, the physiological temperature, have a lower value of backbone entropy compared to any other replica temperature during the REMD simulations. This may indicate a slightly more stable system of sampled structures than what occurs at non-physiological temperatures. This temperature effect on the backbone entropy of the system is not unexpected and is confirmed by circular dichroism studies performed as a function of temperature [4].The sampled secondary structure by the dimer was calculated by the DSSP method [5]. The fraction of sampled secondary structure as a function of residue number in the replica trajectory at 310 K is plotted in Figure 1. The most frequently sampled secondary structure is -turn/-bend followed by -sheet. -Turn/-bend regions...

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supporting

confidence: 89%

Misfolding and Oligomerization of Amyloid β(1-40)

Lovas¹,

Watts²

2015

Peptides 2015, Proceedings of the 24th American Peptide Symposium

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“…Surprisingly, PrP dimers misfolded remarkably easily, invariably following a single pathway leading to the same state. Such homogeneous misfolding contrasts sharply with the heterogeneity seen in SMFS studies of dimers of other aggregation-prone proteins, such as α-synuclein (39) and Aβ (65). The high rate of misfolding also differs starkly from most previous single-molecule studies of tandem-repeat proteins, where misfolding-if it was observed at all-occurred at much lower levels, for example, 2-5% of the time for titin I27 domain repeats (66,67), 4% for tenascin FN III repeats (67), 3-8% for spectrin repeats (68), and 15-30% for α-synuclein repeats (39).…”

Section: Discussionmentioning

confidence: 79%

Protein misfolding occurs by slow diffusion across multiple barriers in a rough energy landscape

Dee

Liu

et al. 2015

Proc. Natl. Acad. Sci. U.S.A.

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The timescale for the microscopic dynamics of proteins during conformational transitions is set by the intrachain diffusion coefficient, D. Despite the central role of protein misfolding and aggregation in many diseases, it has proven challenging to measure D for these processes because of their heterogeneity. We used single-molecule force spectroscopy to overcome these challenges and determine D for misfolding of the prion protein PrP. Observing directly the misfolding of individual dimers into minimal aggregates, we reconstructed the energy landscape governing nonnative structure formation. Remarkably, rather than displaying multiple pathways, as typically expected for aggregation, PrP dimers were funneled into a thermodynamically stable misfolded state along a single pathway containing several intermediates, one of which blocked native folding. Using Kramers' rate theory, D was found to be 1,000-fold slower for misfolding than for native folding, reflecting local roughening of the misfolding landscape, likely due to increased internal friction. The slow diffusion also led to much longer transit times for barrier crossing, allowing transition paths to be observed directly for the first time to our knowledge. These results open a new window onto the microscopic mechanisms governing protein misfolding.intrachain diffusion | protein aggregation | prion protein | optical tweezers | single-molecule force spectroscopy T he formation of intricate 3D structures by proteins is a complex physical process. Such "folding" is typically described in terms of energy landscape theory (1) as a thermally driven diffusive search over an energy landscape in conformational space for the minimum-energy structure. In this picture, whereas the rates at which structural transitions take place are dominated by the presence of energy barriers in the landscape (2), it is the coefficient of diffusion over the landscape, D, that encapsulates the microscopic dynamics of the protein chain, setting the characteristic timescale for molecular motions. Knowledge of D provides insight into the internal friction in the protein chain as it undergoes conformational fluctuations (3) and sets the ultimate speed limit at which changes in structure can take place (4).Given the fundamental importance of the diffusion coefficient in protein folding, there has been much interest in measuring D under different conditions. Conformational diffusion has been studied extensively in peptides and unfolded proteins (5-10), using fluorescence probes such as fluorophore quenching or Förster resonant energy transfer to measure reconfiguration times. Typically, D ∼10 7 -10 8 nm 2 /s was found, although values as low as 10 5 nm 2 /s have been reported (10). Because the diffusion coefficient is inversely proportional to friction, measurements of D have been important for investigating the role and origin of internal friction along the folding pathway (6, 9). Possible links between the value of D and aggregation propensity have also been explored in intrinsically disorde...

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“…Surprisingly, as follow-up studies collected more data on a variety of different molecular systems, a peculiar commonality began to emerge from these experiments: Most showed nonlinear trends in the force spectra. Even a cursory search of the literature finds more than 55 publications that report nonlinear force spectra (see SI Text for details) and cover a large variety of intermolecular systems involving macromolecules (10), peptides (11), small molecules (12), inorganic probes and self-assembled monolayers (13), and binding in both aqueous and organic solvents (14).…”

mentioning

confidence: 99%