Analysis of the Thermodynamics of Binding of an SH3 Domain to Proline-rich Peptides using a Chimeric Fusion Protein

Candel, Adela M.; Nuland, Nico A. J. van; Martin-Sierra, Francisco M.; Martínez, José C.; Conejero‐Lara, Francisco

doi:10.1016/j.jmb.2007.11.060

Cited by 16 publications

(9 citation statements)

References 70 publications

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“…S9B, R 2 = 0.04). This is in agreement with other thermodynamic studies [33] showing that the greater the amount of hydrophobic burial and restrictions in soft motional modes (predominantly in and around SII), the more negative the heat capacity change becomes [34], [35]. AbpSH3:ArkA17 complex formation leads to a ΔC p of −0.36 kcal/mol K which may be accounted for by some combination of the burial of SII, the folding and burial of the C-terminal part of the peptide (all leading to hydrophobic burial) and conformational restrictions in and around SII.…”

Section: Resultssupporting

confidence: 94%

Differential Dynamic Engagement within 24 SH3 Domain: Peptide Complexes Revealed by Co-Linear Chemical Shift Perturbation Analysis

et al. 2012

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There is increasing evidence for the functional importance of multiple dynamically populated states within single proteins. However, peptide binding by protein-protein interaction domains, such as the SH3 domain, has generally been considered to involve the full engagement of peptide to the binding surface with minimal dynamics and simple methods to determine dynamics at the binding surface for multiple related complexes have not been described. We have used NMR spectroscopy combined with isothermal titration calorimetry to comprehensively examine the extent of engagement to the yeast Abp1p SH3 domain for 24 different peptides. Over one quarter of the domain residues display co-linear chemical shift perturbation (CCSP) behavior, in which the position of a given chemical shift in a complex is co-linear with the same chemical shift in the other complexes, providing evidence that each complex exists as a unique dynamic rapidly inter-converting ensemble. The extent the specificity determining sub-surface of AbpSH3 is engaged as judged by CCSP analysis correlates with structural and thermodynamic measurements as well as with functional data, revealing the basis for significant structural and functional diversity amongst the related complexes. Thus, CCSP analysis can distinguish peptide complexes that may appear identical in terms of general structure and percent peptide occupancy but have significant local binding differences across the interface, affecting their ability to transmit conformational change across the domain and resulting in functional differences.

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Section: Resultssupporting

confidence: 94%

Differential Dynamic Engagement within 24 SH3 Domain: Peptide Complexes Revealed by Co-Linear Chemical Shift Perturbation Analysis

et al. 2012

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“…2 and Table 3, the method is successful at purifying both proteins, however, AbpSH3-ArkA protein purifies overall much better than AbpSH3 which may be due to the fact that the peptide binds and protects the domain from unfolding and subsequent aggregation or degradation, consistent with studies from another SH3 domain-peptide hybrid [23]. It should be noted that the band at the same position as AbpSH3-ArkA in the native purification flow through is lysozyme, which is used to lyze the cells in this method but does not have a his-tag and bind to the Ni-NTA columns (data not shown).…”

Section: Notesmentioning

confidence: 62%

“…AbpSH3 has several known peptide targets including a 17-residue peptide from the yeast protein Ark1p [11, 12, 17, 18]. We recently found that AbpSH3 most likely binds in an intra-molecular fashion to a proline rich sequence upstream of the domain [19] and this prompted us to make an AbpSH3-ArkA hybrid to improve expression and purification of the domain, similar to other studies [20–23]. …”

Section: Introductionmentioning

confidence: 60%

A semi-automated method for purification of milligram quantities of proteins on the QIAcube

Mcgraw

Tatipelli

Feyijinmi

et al. 2014

Protein Expression and Purification

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A growing number of studies require the purification of multiple proteins simultaneously and the development of simple economical high-throughput purification methods is essential. We have tested the purification of two related proteins in a variety of conditions to benchmark the semi-automated affinity chromatography method for the QIAcube that we have developed. We find that this new QIAcube method can successfully purify milligram quantities of proteins with minimal user involvement and performs as well as methods based on gravity. The method could easily be adapted to other chromatography resins and should prove to be a versatile method for optimizing protein expression or purification conditions for multiple proteins while obtaining sufficient amounts for subsequent biochemical analyses.

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“…In 2008, we identified more than 500 articles after searching Web of Science, PubMed, SciDir and OVID databases using ‘isothermal AND titration AND calorimetry’ or ITC or ‘Isothermal Titration Calorimetry’ search terms. Following the format of previous annual surveys these have been classified into the following categories: References cited in the text and review articles 1–82. Protein–protein and protein–peptide interactions 83–222. Protein–small ligand or protein–drug interactions 223–321. Protein/peptide metal interactions 322–355. Protein/peptide nucleic acid interactions 356–387. Protein/peptide lipid interactions …”

Section: Introductionmentioning

confidence: 99%

Survey of the year 2008: applications of isothermal titration calorimetry

Falconer

Penkova

Jelesarov

et al. 2010

J of Molecular Recognition

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Isothermal titration calorimetry (ITC) is a fast, accurate and label-free method for measuring the thermodynamics and binding affinities of molecular associations in solution. Because the method will measure any reaction that results in a heat change, it is applicable to many different fields of research from biomolecular science, to drug design and materials engineering, and can be used to measure binding events between essentially any type of biological or chemical ligand. ITC is the only method that can directly measure binding energetics including Gibbs free energy, enthalpy, entropy and heat capacity changes. Not only binding thermodynamics but also catalytic reactions, conformational rearrangements, changes in protonation and molecular dissociations can be readily quantified by performing only a small number of ITC experiments. In this review, we highlight some of the particularly interesting reports from 2008 employing ITC, with a particular focus on protein interactions with other proteins, nucleic acids, lipids and drugs. As is tradition in these reviews we have not attempted a comprehensive analysis of all 500 papers using ITC, but emphasize those reports that particularly captured our interest and that included more thorough discussions we consider exemplify the power of the technique and might serve to inspire other users.

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Analysis of the Thermodynamics of Binding of an SH3 Domain to Proline-rich Peptides using a Chimeric Fusion Protein

Cited by 16 publications

References 70 publications

Differential Dynamic Engagement within 24 SH3 Domain: Peptide Complexes Revealed by Co-Linear Chemical Shift Perturbation Analysis

Differential Dynamic Engagement within 24 SH3 Domain: Peptide Complexes Revealed by Co-Linear Chemical Shift Perturbation Analysis

A semi-automated method for purification of milligram quantities of proteins on the QIAcube

Survey of the year 2008: applications of isothermal titration calorimetry

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