A native to amyloidogenic transition regulated by a backbone trigger

Eakin, Catherine M.; Berman, Andrea J.; Miranker, Andrew D.

doi:10.1038/nsmb1068

Cited by 184 publications

(294 citation statements)

References 57 publications

Supporting

Mentioning

271

Contrasting

Order By: Relevance

“…16 10 We therefore employed mutagenesis to generate a more uniform population. Mutations were targeted based on predicted interfaces 20 and a single point mutant, H13F, was sufficient to increase both the extent and homogeneity of the oligomerization reaction, resulting in efficient formation of a hexamer. Nevertheless, the global stability, Cu 2+ affinity, and rate of oligomerization remain similar for H13F and WT β2m.…”

Section: Cu 2+ Mediated Oligomerizationmentioning

confidence: 99%

“…This led to the suggestion that oligomerization is rate limited by a conformational rearrangement from the native to an 'activated' monomeric state which we termed 'M*'. 10,20 Another state, termed I T , identified by refolding studies may be related to this structure. 6,21 Oligomers are native-like by circular dichroism and 1 H NMR yet at the same time exhibit characteristics of amyloid as evidenced by the fluorescence response of a histological dye, Thioflavin T. 22 This supports a model for β2m oligomers where the global fold of β2m is maintained.…”

Section: Cu 2+ Mediated Oligomerizationmentioning

confidence: 99%

See 1 more Smart Citation

Metal binding sheds light on mechanisms of amyloid assembly

Calabrese

Miranker

2009

Prion

View full text Add to dashboard Cite

β-2 microglobulin (β2m) is the protein responsible for amyloid deposition in Dialysis-Related Amyloidosis (DRA). Aggregation can be induced by various solution conditions including exposure to divalent metal, incubation at acidic pH, and limited proteolysis. Using Cu 2+ as a trigger, we have trapped, isolated, and crystallized a stable oligomer of β2m that is populated under amyloidogenic solution conditions (Calabrese et al. Nat Struct Mol Biol 2008; 15:965-71). This structure reveals that Cu 2+ -binding is associated with dramatic conformational rearrangements. This has allowed us to postulate a set of structural changes common to all β2m aggregation pathways. Cu 2+ serves as a potential trigger in other aggregation systems such as Aβ, α-synuclein, and mammalian Prion (PrP). A comparison of Cu 2+ binding to β2m and PrP reveals common features. Therefore, in addition to providing insight into DRA, induction of structure by Cu 2+ binding appears to be a recurring structural motif for pathological changes in conformation. Backgroundβ2m is the 99 residue, 12 kDa non-covalent light chain of the Class I Major Histocompatibility Complex (MHC). Its function is to ensure proper folding and cell-surface expression of the MHC. 1 In the course of normal turnover, β2m is released to the serum and catabolized by the kidneys. In patients suffering from endstage renal disease who are treated with hemodialysis therapy, β2m levels rise and amyloid plaques develop throughout the joints and connective tissue. 2 Although an elevated serum level of β2m may be necessary for aggregation, it is not sufficient as β2m remains stably folded at > 1 mM concentration at 37 °C. 3,4 Furthermore, it can be reversibly folded in vitro 5-7 and amyloid is not reported in other diseases with elevated levels in serum. 8,9 Therefore, aggregation must be triggered by features unique to hemodialysis therapy.As β2m exists as a stable monomer under near physiological conditions, 3,10 much effort has been focused on understanding the molecular mechanism by which self association is triggered. Many approaches have been used to induce β2m aggregation. These include limited proteolysis, 11 incubation at acidic pH, 12 and exposure to detergents or organic solvents. 13,14 In 2001, we discovered that β2m is a Cu 2+ -binding protein. 4 Whereas β2m apo is not amyloidogenic, β2m holo is aggregation-prone. Cu 2+ Mediated OligomerizationWe are interested in Cu 2+ -dependent aggregation of β2m for several reasons. First, hemodialysis patients may be exposed to elevated levels of divalent cation as a consequence of therapy. This is apparent in the historical use of Cu 2+ in the preparation of dialysis membranes, and in water quality standards which permit as much as 1.6 μM Cu 2+ in hemodialysate. 15 This is comparable to the ~3 μM affinity of β2m measured in vitro. 4 Second, early experiments revealed that although Cu 2+ is necessary to initiate aggregation, mature fibers remain stable in the presence of a metal chelate. 10 This allowed the possibility that Cu 2+ acti...

show abstract

Section: Cu 2+ Mediated Oligomerizationmentioning

confidence: 99%

Section: Cu 2+ Mediated Oligomerizationmentioning

confidence: 99%

Metal binding sheds light on mechanisms of amyloid assembly

Calabrese

Miranker

2009

Prion

View full text Add to dashboard Cite

show abstract

“…All NMR spectra were acquired on a Bruker DRX700 spectrometer (Bruker Corp., Germany) equipped with 5 mm triple resonance probe on peptide samples of ~1 mM concentration at 6.5 < pH < 7.1 (10% D 2 O and DSS as the internal proton reference standard set to 0 ppm) at 288 K. 1 Transmission electron microscopy. Samples were diluted 20-fold with water and applied to 300-mesh formvar/carbon-coated copper grids, allowed to adhere for 1 min, and stained for 40 sec with 1% uranyl acetate.…”

Section: Methodsmentioning

confidence: 99%

“…1 Apart from the disease-related amyloids, some functional amyloid proteins has also been discovered which play an important role e.g. in melanin synthesis or in bacterial biofilm formation.…”

Section: Introductionmentioning

confidence: 99%

Phosphorylation as Conformational Switch from the Native to Amyloid State: Trp-Cage as a Protein Aggregation Model

et al. 2015

View full text Add to dashboard Cite

The 20 residue long Trp-cage miniprotein is an excellent model both for computational and experimental studies of protein folding and stability. Recently, a great attention emerged to study disease-related protein misfolding, aggregation and amyloid formation, with the aim of revealing their structural and thermodynamic background. Trp-cage is sensitive to both environmental and structure-modifying effects, and aggregates with ease upon structure destabilization and thus, it is an ideal model of aggregation and amyloid formation. Here, we characterize the amyloid formation of several sequence modified and side-chain phosphorylated Trp-cage variants. We applied NMR, CD, FTIR spectroscopy, MD simulations, transmission electron microscopy in conjunction with thioflavin-T fluorescence measurements to reveal the structural consequences of side-chain phosphorylation. We demonstrate that the native fold is destabilized upon serine phosphorylation and the resultant highly dynamic structures form amyloid-like ordered aggregates with high intermolecular β-structure content. The only exception is the D9S(P) variant which follows an alternative aggregation process by forming thin fibrils, presenting a CD spectrum of PPII helix, and showing low ThT binding capability. We propose a complex aggregation model for these Trp-cage miniproteins. This model assumes an additional aggregated state, a collagen triple helical form which can precede amyloid formation. The phosphorylation of a single serine residue serves as a conformational switch, triggering aggregation, otherwise mediated by kinases in cell. We show that Trp-cage miniprotein is indeed a realistic model of larger globular systems of composite folding and aggregation landscapes and helps us to understand the fundamentals of deleterious protein aggregation and amyloid formation.3

show abstract

“…The alteration in the hydrogen bond interactions lead to the increase in the flexibility of the residues. 48 Thus, hydrogen bond analysis in the loop regions was performed to investigate the enhanced flexibility of the loop regions in D59P as compare to wt β2m.…”

Section: Hydrogen Bond Analysis In the Loop Regionsmentioning

confidence: 99%

Assessing the effect of D59P mutation in the DE loop region in amyloid aggregation propensity of β2‐microglobulin: A molecular dynamics simulation study

Narang

Shuaib

Goyal

2017

J of Cellular Biochemistry

View full text Add to dashboard Cite

Dialysis-related amyloidosis (DRA) is a severe condition characterized by the accumulation of amyloidogenic β2-microglobulin (β2m) protein around skeletal joints and bones. The recent studies highlighted a critical role of the DE loop region for β2m stability and amyloid aggregation propensity. Despite significant efforts, the molecular mechanism of enhanced aggregation due to D59P mutation in the DE loop region remain elusive. In the present study, explicit-solvent molecular dynamics (MD) simulations were performed to examine the key changes in the structural and dynamic properties of wild type (wt) β2m upon D59P mutation. MD simulations reveal a decrease in the average number of hydrogen bonds in the loop regions on D59P mutation that enhances conformational flexibility, which lead to higher aggregation propensity of D59P as compare to wt β2m. The principal component analysis (PCA) highlight that D59P covers a larger region of phase space and display a higher trace value than wt β2m, which suggest an overall enhancement in the conformational flexibility. D59P display two minimum energy basins in the free energy landscape (FEL) that are associated with thermodynamically less stable conformational states as compare to single minimum energy basin in wt β2m. The present study provides theoretical insights into the molecular mechanism behind the higher aggregation propensity of D59P as compare to wt β2m. K E Y W O R D Samyloid aggregation, β2-microglobulin, D59P mutation, DE loop, molecular dynamics

show abstract

A native to amyloidogenic transition regulated by a backbone trigger

Cited by 184 publications

References 57 publications

Metal binding sheds light on mechanisms of amyloid assembly

Metal binding sheds light on mechanisms of amyloid assembly

Phosphorylation as Conformational Switch from the Native to Amyloid State: Trp-Cage as a Protein Aggregation Model

Assessing the effect of D59P mutation in the DE loop region in amyloid aggregation propensity of β2‐microglobulin: A molecular dynamics simulation study

Contact Info

Product

Resources

About