Probing transient non-native states in amyloid beta fiber elongation by NMR

Brender, Jeffrey R.; Ghosh, Anirban; Kotler, Samuel A.; Krishnamoorthy, Janarthanan; Bera, Swapna; Morris, Vanessa K.; Sil, Timir Baran; Garai, Kanchan; Reif, Bernd; Bhunia, Anirban; Ramamoorthy, Ayyalusamy

doi:10.1039/c9cc01067j

Cited by 47 publications

(56 citation statements)

References 29 publications

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“…In this context, it is indispensable to discuss the far-reaching possibilities of NMR experiments to enable the indirect probing of the nucleation events in the amyloidogenic pathways. 33,55 The protein-folding intermediates have been implicated in harbouring the essential chemical cues that provide useful insight into understanding the amyloidogenic propensity. 33,55 These intermediates are often structurally similar to the low molecular weight early conformers of the protein but form a chemically distinct pool that nucleates the early aggregation events.…”

Section: Discussionmentioning

confidence: 99%

“…33,55 The protein-folding intermediates have been implicated in harbouring the essential chemical cues that provide useful insight into understanding the amyloidogenic propensity. 33,55 These intermediates are often structurally similar to the low molecular weight early conformers of the protein but form a chemically distinct pool that nucleates the early aggregation events. High-resolution NMR spectroscopy has helped us to unveil the mechanism of Aβ amyloidogenesis in Alzheimer's disease to be actually a multi-step process involving the docking of the free monomeric forms to the nucleating intermediates.…”

Section: Discussionmentioning

confidence: 99%

“…High-resolution NMR spectroscopy has helped us to unveil the mechanism of Aβ amyloidogenesis in Alzheimer's disease to be actually a multi-step process involving the docking of the free monomeric forms to the nucleating intermediates. 55,56 Similarly, in Parkinson's disease, atomic-resolution characterization of the αsynuclein protein enabled us to probe the "open" conformation of the monomeric form that serves as the intermediate in initiating the amyloidogenic cascade. 56 Previous studies have also indicated that the amyloid fibril formation for globular proteins, such as HI, occurs via partially unfolded intermediates that gradually associate to form the oligomers, eventually into well-ordered mature fibrils.…”

Section: Discussionmentioning

confidence: 99%

“…9,14,23,27,33,47,57,58 A partial unfolding of the monomeric globular protein produces a molten globule like structure that is comparable to the amyloidogenic protein intermediates, acting as the nucleating form. 29,50,55,[59][60][61] In the absence of zinc at physiological pH, HI is primarily present as a conformationally stable dimer;…”

Section: Discussionmentioning

confidence: 99%

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Mechanistic Studies of the Stabilization of Insulin Helical Structure by Coomassie Brilliant Blue

Dolui¹,

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Saha³

et al. 2020

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Human insulin (HI) is an essential protein hormone and its biological activity mostly depends on folded and active conformation in the monomeric state. The present investigation established that Coomassie Brilliant Blue G-250 (CBBG), a small multicyclic hydroxyl compound can reversibly bind to the hormonal protein dimer and maintained most of α-helical folds crucial for biological function of the enzyme. The solution-state 1D NMR and isothermal calorimetric analysis showed a sub-micromolar binding affinity of the molecule to HI. 2D NOESY NMR established that the HI dimer undergoes residue level local conformational change upon binding to CBBG. The chemical shift perturbation and the NOE parameters of active protons of amino acid residues throughout the polypeptides further suggested that CBBG upon binding the protein stabilize α-helixes of both the A and B subunits of the hormonal protein. The changes in Gibb’s free energy (∆G) of the binding was of ~−11.1 kcal/mol and suggested a thermodynamically favourable process. The changes in enthalpy (∆H) and entropy term (T∆S) were −57.2 kcal/mol and −46.1 kcal/mol, respectively. The negative changes in entropy and the NOE transfer effectiveness of several residues in the presence of CBBG molecules indicated that the binding was an enthalpy driven favourable equilibrium process. The NMR-based atomic resolution data and molecular docking studies confirmed that the CBBG binds to HI at the dimeric stage and prevents the availability of the crucial residue segments that partake directly in further oligomerization and subsequent fibrillation. Extended computational analysis based on chemical shift perturbation of protons of active residues further established receptor-ligand based pharmacophore model comprised of 5 hydrophobic and a hydrogen bond acceptor features that can anchor the residues at the A and B chains of HI and inhibit the partial unfolding and hydrophobic collapse to nucleate the fibrillation. Taken together, the results demonstrated that CBBG and their close analogues might be useful to develop a formulation that will maintain the active and functional form of the hormonal protein for a significantly longer time.TOC

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Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

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Mechanistic Studies of the Stabilization of Insulin Helical Structure by Coomassie Brilliant Blue

Dolui¹,

Pariary

Saha³

et al. 2020

Preprint

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“…Instead, they likely arise from smaller β-sheet containing oligomers, possibly catalyzed by interaction with the fiber surface. [61][62][63] While we could not make a definitive assignment due to the lack of i,i+2 NOEs, it is possible to check the consistency of the two new NOE peaks with the model of the insulin amyloid fiber proposed by Ivanova et al Both H10 B and V18 B and V12 B and Y16 B are in close in contact with each other in the model in which the the helical region of B chain forms anti-parallel β-sheets along the fiber axis ( Fig. S4).…”

Section: A Late Stage Intermediate Contains An Anti-parallel β-Sheetmentioning

confidence: 99%

High resolution structure of a partially folded insulin aggregation intermediate

Ratha

Kar

Brender

et al. 2019

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Insulin has long served as a model for protein aggregation, both because of the importance of aggregation in insulin manufacture and because the structural biology of insulin has been extensively characterized. Despite intensive study, details about the initial triggers for aggregation have remained elusive at the molecular level. We show here that at acidic pH, the aggregation of insulin is likely initiated by a partially folded monomeric intermediate whose concentration is controlled by an off-pathway micellar species. High resolution structures of the partially folded intermediate show that it is coarsely similar to the initial monomeric structure but differs in subtle details -the A chain helices on the receptor interface are more disordered and the B chain helix moves away from C-terminal A chain helix. The result of these movements is the creation of a hydrophobic cavity in the center of the protein that may serve as nucleation site for oligomer formation. Knowledge of this transition may aid in the engineering of insulin variants that retain the favorable pharamacokinetic properties of monomeric insulin but are more resistant to aggregation.

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Self‐Assembly and Neurotoxicity of β‐Amyloid (21–40) Peptide Fragment: The Regulatory Role of GxxxG Motifs

et al. 2019

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The three GxxxG repeating motifs from the C-terminal region of β-amyloid (Aβ) peptide play a significant role in regulating the aggregation kinetics of the peptide. Mutation of these glycine residues to leucine greatly accelerates the fibrillation process but generates a varied toxicity profile. Using an array of biophysical techniques, we demonstrated the uniqueness of the composite glycine residues in these structural repeats. We used solvent relaxation NMR spectroscopy to investigate the role played by the surrounding water molecules in determining the corresponding aggregation pathway. Notably, the conformational changes induced by Gly 33 and Gly 37 mutations result in significantly decreased toxicity in a neuronal cell line. Our results indicate that G 33 xxxG 37 is the primary motif responsible for Aβ neurotoxicity, hence providing a direct structurefunction correlation. Targeting this motif, therefore, can be a promising strategy to prevent neuronal cell death associated with Alzheimer's and other related diseases, such as type II diabetes and Parkinson's. ROESY were 48 and 64, respectively. Data processing and analysis were carried out using Topspin TM v3.2 software (Bruker Biospin, Switzerland) and Sparky (https://www.cgl.ucsf.edu/home/sparky) software, respectively.

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Probing transient non-native states in amyloid beta fiber elongation by NMR

Abstract: Using NMR to probe transient binding of Aβ1–40 monomers to fibers, we find partially bound conformations with the highest degree of interaction near F19–K28 and a lesser degree of interaction near the C-terminus (L34–G37).

Cited by 47 publications

References 29 publications

Mechanistic Studies of the Stabilization of Insulin Helical Structure by Coomassie Brilliant Blue

Mechanistic Studies of the Stabilization of Insulin Helical Structure by Coomassie Brilliant Blue

High resolution structure of a partially folded insulin aggregation intermediate

Self‐Assembly and Neurotoxicity of β‐Amyloid (21–40) Peptide Fragment: The Regulatory Role of GxxxG Motifs

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