Circular Dichroism Techniques for the Analysis of Intrinsically Disordered Proteins and Domains

2012 ◽  
pp. 387-404 ◽  
Author(s):  
Lucía B. Chemes ◽  
Leonardo G. Alonso ◽  
María G. Noval ◽  
Gonzalo de Prat-Gay
Keyword(s):  
Circular Dichroism ◽  
Intrinsically Disordered Proteins ◽  
Disordered Proteins ◽  
Intrinsically Disordered ◽  
Circular Dichroïsm

scholarly journals Helix formation during the coupled binding and folding of intrinsically disordered proteins monitored by synchrotron-radiation circular dichroism spectroscopy

2019 ◽  
Author(s):  
Elin Karlsson ◽  
Eva Andersson ◽  
Nykola C. Jones ◽  
Søren Vrønning Hoffmann ◽  
Per Jemth ◽  
...  
Keyword(s):  
Circular Dichroism ◽  
Ionic Strength ◽  
Synchrotron Radiation ◽  
Intrinsically Disordered Proteins ◽  
Circular Dichroism Spectroscopy ◽  
High Ionic Strength ◽  
Disordered Proteins ◽  
Intrinsically Disordered ◽  
Helix Formation ◽  
Circular Dichroïsm

AbstractIntrinsically disordered proteins organize interaction networks in the cell in many regulation and signalling processes. These proteins often gain structure upon binding to their target proteins in multi-step reactions involving the formation of both secondary and tertiary structure. To understand the interactions of disordered proteins, we need to understand the mechanisms of these coupled folding and binding reactions. We studied helix formation in the binding of the molten globule-like nuclear coactivator binding domain (NCBD) and the disordered interaction domain from activator of thyroid hormone and retinoid receptors (ACTR). We demonstrate that helix formation in a rapid binding reaction can be followed by stopped flow synchrotron-radiation circular dichroism spectroscopy, and describe the design of such a beamline. Fluorescence-monitored binding experiments of ACTR and NCBD display several kinetic phases including one concentration-independent phase, which is consistent with an intermediate stabilized at high ionic strength. Time resolved circular dichroism experiments show that almost all helicity is formed upon initial association of the proteins, or separated from the encounter complex by only a small energy barrier. Through simulation of mechanistic models, we show that the intermediate observed at high ionic strength likely involves a structural rearrangement with minor overall changes in helicity. Our experiments provide a benchmark for simulations of coupled binding reactions and demonstrate the feasibility of using synchrotron radiation circular dichroism for mechanistic studies of protein-protein interactions.

scholarly journals SESCA: Predicting Circular Dichroism Spectra from Protein Molecular Structures

2018 ◽  
Author(s):  
Gabor Nagy ◽  
Maxim Igaev ◽  
Søren V. Hoffmann ◽  
Nykola C. Jones ◽  
Helmut Grubmüller
Keyword(s):  
Circular Dichroism ◽  
Secondary Structure ◽  
Disordered Proteins ◽  
Basis Sets ◽  
Cd Spectra ◽  
Circular Dichroism Spectra ◽  
Essential Information ◽  
Intrinsically Disordered ◽  
Circular Dichroïsm ◽  
Secondary Structure Composition

AbstractCircular dichroism spectroscopy is a highly sensitive, but low-resolution technique to study the structure of proteins. Combined with molecular modelling or other complementary techniques, CD spectroscopy can provide essential information at higher resolution. To this end, we introduce a new computational method to calculate the electronic circular dichroism spectra of proteins from a structural model or ensemble using the average secondary structure composition and a pre-calculated set of basis spectra. We compared the predictive power of our method to existing algorithms – namely DichroCalc and PDB2CD – and found that it predicts CD spectra more accurately, with a 50% smaller average deviation from the measured CD spectra. Our results indicate that the derived basis sets are robust to experimental errors in the reference spectra and to the choice of the secondary structure classification algorithm. For over 80% of the globular reference proteins, our basis sets accurately predict the experimental spectrum solely from their secondary structure composition. For the remaining 20%, correcting for intensity normalization considerably improves the prediction power. Additionally, we show that the predictions for short peptides and intrinsically disordered proteins strongly benefit from accounting for side-chain contributions and structural flexibility.Table Of Content Graphics:

Sequence Specificity Despite Intrinsic Disorder: How a Disease-Associated Val/Met Polymorphism Shifts Tertiary Interactions in a Long Disordered Protein

2019 ◽  
Author(s):  
Ruchi Lohia ◽  
Reza Salari ◽  
Grace Brannigan
Keyword(s):  
Secondary Structure ◽  
Electrostatic Interactions ◽  
Intrinsically Disordered Proteins ◽  
Disordered Proteins ◽  
Sequence Specificity ◽  
Intrinsically Disordered ◽  
Specific Effects ◽  
Heterogeneous Polymer ◽  
Non Local ◽  
Dynamics Simulations

<div>The role of electrostatic interactions and mutations that change charge states in intrinsically disordered proteins (IDPs) is well-established, but many disease-associated mutations in IDPs are charge-neutral. The Val66Met single nucleotide polymorphism (SNP) encodes a hydrophobic-to-hydrophobic mutation at the midpoint of the prodomain of precursor brain-derived neurotrophic factor (BDNF), one of the earliest SNPs to be associated with neuropsychiatric disorders, for which the underlying molecular mechanism is unknown. Here we report on over 250 μs of fully-atomistic, explicit solvent, temperature replica exchange molecular dynamics simulations of the 91 residue BDNF prodomain, for both the V66 and M66 sequence.</div><div>The simulations were able to correctly reproduce the location of both local and non-local secondary changes due to the Val66Met mutation when compared with NMR spectroscopy. We find that the local structure change is mediated via entropic and sequence specific effects. We show that the highly disordered prodomain can be meaningfully divided into domains based on sequence alone. Monte Carlo simulations of a self-excluding heterogeneous polymer, with monomers representing each domain, suggest the sequence would be effectively segmented by the long, highly disordered polyampholyte near the sequence midpoint. This is qualitatively consistent with observed interdomain contacts within the BDNF prodomain, although contacts between the two segments are enriched relative to the self-excluding polymer. The Val66Met mutation increases interactions across the boundary between the two segments, due in part to a specific Met-Met interaction with a Methionine in the other segment. This effect propagates to cause the non-local change in secondary structure around the second methionine, previously observed in NMR. The effect is not mediated simply via changes in inter-domain contacts but is also dependent on secondary structure formation around residue 66, indicating a mechanism for secondary structure coupling in disordered proteins. </div>

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