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Structural characterization of Hsp12, the heat shock protein from Saccharomyces cerevisiae, in aqueous solution where it is intrinsically disordered and in detergent micelles where it is locally ?-helical.

ABSTRACT: Hsp12 (heat shock protein 12) belongs to the small heat shock protein family, partially characterized as a stress response, stationary phase entry, late embryonic abundant-like protein located at the plasma membrane to protect membrane from desiccation. Here, we report the structural characterization of Hsp12 by NMR and biophysical techniques. The protein was labeled uniformly with nitrogen-15 and carbon-13 so that its conformation could be determined in detail both in aqueous solution and in two membrane-mimetic environments, SDS and dodecylphosphocholine (DPC) micelles. Secondary structural elements determined from assigned chemical shifts indicated that Hsp12 is dynamically disordered in aqueous solution, whereas it gains four helical stretches in the presence of SDS micelles and a single helix in presence of DPC. These conclusions were reinforced by circular dichroism spectra of the protein in all three environments. The lack of long range interactions in NOESY spectra indicated that the helices present in SDS micelles do not pack together. R(1) and R(2), relaxation and heteronuclear NOE measurements showed that the protein is disordered in aqueous solution but becomes more ordered in presence of detergent micelles. NMR spectra collected in presence of paramagnetic spin relaxation agents (5DSA, 16DSA, and Gd(DTPA-BMA)) indicated that the amphipathic ?-helices of Hsp12 in SDS micelles lie on the membrane surface. These observations are in agreement with studies suggesting that Hsp12 functions to protect the membrane from desiccation.

SUBMITTER: Singarapu KK

PROVIDER: S-EPMC3234819 | biostudies-literature | 2011 Dec

REPOSITORIES: biostudies-literature

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Structural characterization of Hsp12, the heat shock protein from Saccharomyces cerevisiae, in aqueous solution where it is intrinsically disordered and in detergent micelles where it is locally α-helical.

Singarapu Kiran K KK Tonelli Marco M Chow Darius C DC Frederick Ronnie O RO Westler William M WM Markley John L JL

The Journal of biological chemistry 20111013 50

Hsp12 (heat shock protein 12) belongs to the small heat shock protein family, partially characterized as a stress response, stationary phase entry, late embryonic abundant-like protein located at the plasma membrane to protect membrane from desiccation. Here, we report the structural characterization of Hsp12 by NMR and biophysical techniques. The protein was labeled uniformly with nitrogen-15 and carbon-13 so that its conformation could be determined in detail both in aqueous solution and in tw ...[more]

PMID: 21998307

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Project description:Intrinsically disordered proteins (IDPs) are abundant in eukaryotic proteomes and preform critical roles in many cellular processes, most often through the association with globular proteins. Despite lacking a stable three-dimensional structure by themselves, they may acquire a defined conformation upon binding globular targets. The most common type of secondary structure acquired by these binding motifs entails formation of an α-helix. It has been hypothesized that such disorder-to-order transitions are associated with a significant free energy penalty due to IDP folding, which reduces the overall IDP-target affinity. However, the exact magnitude of IDP folding penalty in α-helical binding motifs has not been systematically estimated. Here, we report the folding penalty contributions for 30 IDPs undergoing folding-upon-binding and find that the average IDP folding penalty is +2.0 kcal/mol and ranges from 0.7 to 3.5 kcal/mol. We observe that the folding penalty scales approximately linearly with the change in IDP helicity upon binding, which provides a simple empirical way to estimate folding penalty. We analyze to what extent do pre-structuring and target-bound IDP dynamics (fuzziness) reduce the folding penalty and find that these effects combined, on average, reduce the folding cost by around half. Taken together, the presented analysis provides a quantitative basis for understanding the role of folding penalty in IDP-target interactions and introduces a method estimate this quantity. Estimation and reduction of IDP folding penalty may prove useful in the rational design of helix-stabilized inhibitors of IDP-target interactions. STATEMENT: The α-helical binding motifs are ubiquitous among the intrinsically disordered proteins (IDPs). Upon binding their targets, they undergo a disorder-to-order transition, which is accompanied by a significant folding penalty whose magnitude is generally not known. Here, we use recently developed statistical-thermodynamic model to estimate the folding penalties for 30 IDPs and clarify the roles of IDP pre-folding and bound-state dynamics in reducing the folding penalty.

Dataset Information

Structural characterization of Hsp12, the heat shock protein from Saccharomyces cerevisiae, in aqueous solution where it is intrinsically disordered and in detergent micelles where it is locally ?-helical.

Publications

Structural characterization of Hsp12, the heat shock protein from Saccharomyces cerevisiae, in aqueous solution where it is intrinsically disordered and in detergent micelles where it is locally α-helical.

Similar Datasets

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