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Dissecting the Energetics of Intrinsically Disordered Proteins via a Hybrid Experimental and Computational Approach.

ABSTRACT: Intrinsically disordered proteins (IDPs) play important roles in biology, but little is known about the energetics of their inter-residue interactions. Methods that have been successfully applied to analyze the energetics of globular proteins are not applicable to the fluctuating partially ordered ensembles populated by IDPs. A combined computational experimental strategy is introduced for analyzing the energetic role of individual residues in the free state of IDPs. The approach combines experimental measurements of the binding of wild-type and mutant IDPs to their partners with alchemical free energy calculations of the structured complexes. These data allow quantitative information to be deduced about the free state via a thermodynamic cycle. The approach is validated by the analysis of the effects of mutations upon the binding free energy of the ovomucoid inhibitor third binding domain to its partners and is applied to the C-terminal domain of the measles virus nucleoprotein, a 125-residue IDP involved in the RNA transcription and replication of measles virus. The analysis reveals significant inter-residue interactions in the unbound IDP and suggests a biological role for them. The work demonstrates that advances in force fields and computational hardware have now led to the point where it is possible to develop methods, which integrate experimental and computational techniques to reveal insights that cannot be studied using either technique alone.

SUBMITTER: Zou J

PROVIDER: S-EPMC7291390 | biostudies-literature | 2019 Dec

REPOSITORIES: biostudies-literature

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Dissecting the Energetics of Intrinsically Disordered Proteins via a Hybrid Experimental and Computational Approach.

Zou Junjie J Simmerling Carlos C Raleigh Daniel P DP

The journal of physical chemistry. B 20191203 49

Intrinsically disordered proteins (IDPs) play important roles in biology, but little is known about the energetics of their inter-residue interactions. Methods that have been successfully applied to analyze the energetics of globular proteins are not applicable to the fluctuating partially ordered ensembles populated by IDPs. A combined computational experimental strategy is introduced for analyzing the energetic role of individual residues in the free state of IDPs. The approach combines experi ...[more]

PMID: 31702919

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Project description:The broad family of LEA proteins are intrinsically disordered proteins (IDPs) with several potential roles in desiccation tolerance, or anhydrobiosis, one of which is to limit desiccation-induced aggregation of cellular proteins. We show here that this activity, termed molecular shield function, is distinct from that of a classical molecular chaperone, such as HSP70 - while HSP70 reduces aggregation of citrate synthase (CS) on heating, two LEA proteins, a nematode group 3 protein, AavLEA1, and a plant group 1 protein, Em, do not; conversely, the LEA proteins reduce CS aggregation on desiccation, while HSP70 lacks this ability. There are also differences in interaction with client proteins - HSP70 can be co-immunoprecipitated with a polyglutamine-containing client, consistent with tight complex formation, whereas the LEA proteins can not, although a loose interaction is observed by Förster resonance energy transfer. In a further exploration of molecular shield function, we demonstrate that synthetic polysaccharides, like LEA proteins, are able to reduce desiccation-induced aggregation of a water-soluble proteome, consistent with a steric interference model of anti-aggregation activity. If molecular shields operate by reducing intermolecular cohesion rates, they should not protect against intramolecular protein damage. This was tested using the monomeric red fluorescent protein, mCherry, which does not undergo aggregation on drying, but the absorbance and emission spectra of its intrinsic fluorophore are dramatically reduced, indicative of intramolecular conformational changes. As expected, these changes are not prevented by AavLEA1, except for a slight protection at high molar ratios, and an AavLEA1-mCherry fusion protein is damaged to the same extent as mCherry alone. A recent hypothesis proposed that proteomes from desiccation-tolerant species contain a higher degree of disorder than intolerant examples, and that this might provide greater intrinsic stability, but a bioinformatics survey does not support this, since there are no significant differences in the degree of disorder between desiccation tolerant and intolerant species. It seems clear therefore that molecular shield function is largely an intermolecular activity implemented by specialist IDPs, distinct from molecular chaperones, but with a role in proteostasis.

Dataset Information

Dissecting the Energetics of Intrinsically Disordered Proteins via a Hybrid Experimental and Computational Approach.

Publications

Dissecting the Energetics of Intrinsically Disordered Proteins via a Hybrid Experimental and Computational Approach.

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