Modeling the Early Stages of Phase Separation in Disordered Elastin-like Proteins

Zhang, Yue; Zai-Rose, Valeria; Price, C. J.; Ezzell, Nicholas A.; Bidwell, Gene L.; Correia, John J.; Fitzkee, Nicholas C.

doi:10.1016/j.bpj.2018.01.045

Cited by 18 publications

(28 citation statements)

References 96 publications

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“…provided an experimental structural model for the study of the early stages of the phase separation behavior [38]. A 40-pentapeptide ELR suitable for nuclear resonance measurements was developed based on a polyVPGVG that contained a different guest residue every seven pentapeptides (Lys, Thr, Ala, Ile, Ser and Leu).…”

Section: Elrs As Model Intrinsically Disordered Proteinsmentioning

confidence: 99%

Trends in the design and use of elastin-like recombinamers as biomaterials

et al. 2019

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Elastin-like recombinamers (ELRs), which derive from one of the repetitive domains found in natural elastin, have been intensively studied in the last few years from several points of view. In this mini review, we discuss all the recent works related to the investigation of ELRs, starting with those that define these polypeptides as model intrinsically disordered proteins or regions (IDPs or IDRs) and its relevance for some biomedical applications. Furthermore, we summarize the current knowledge on the development of drug, vaccine and gene delivery systems based on ELRs, while also emphasizing the multiple tissue engineering approaches involving their use. Finally, we show different studies that explore applications in other fields, and several examples that 2 describe biomaterial blends in which ELRs have a key role. This review aims to give an overview of the recent advances regarding ELRs and to encourage further investigation of their properties and applications.

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Section: Elrs As Model Intrinsically Disordered Proteinsmentioning

confidence: 99%

Trends in the design and use of elastin-like recombinamers as biomaterials

et al. 2019

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“…The basic CPP constructs associated slightly better at higher temperature, 30-35 C, suggesting a coupling to b-turn formation and an association mechanism involving b-turn-b-turn interactions (see Fig. 2 in (15)). DSC measurements verified for all constructs that the liquid-liquid phase separation (LLPS) is driven by a favorable and positive entropy (DS) consistent with the hydrophobic effect, the loss of water (''desolvation''), and the burial of hydrophobic surface (''collapse'').…”

Section: Introductionmentioning

confidence: 96%

“…To test some of these predictions, ensembles of ELP1 structures with increasing amounts of b-turns were generated by structure-biased Monte Carlo simulations (16,17) and compared to experimental hydrodynamic data on mono-meric and dimeric ELP1 species (15). Dimers were simulated assuming that b-turns mediated dimer stability in a modular manner.…”

Section: Introductionmentioning

confidence: 99%

“…Buried hydrophobic surface is the predominant driving force for the hydrophobic effect observed experimentally with ELP. To further test these results, NMR experiments were performed on a short ELP construct (ELP40; see Table 1) with six nonvalyl guest residues to allow identification of structural signatures throughout the sequence (15). Our results and recent NMR experiments (18) and molecular dynamics simulations (19) confirm the presence of transient b-turns (20-40% by NMR (18) and 16-20% by molecular dynamics (19)), and it is these turns that we hypothesize are involved in intermolecular hydrophobic contacts.…”

Section: Introductionmentioning

confidence: 99%

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Effects of Doxorubicin on the Liquid-Liquid Phase Change Properties of Elastin-Like Polypeptides

Zai-Rose

West

Krämer

et al. 2018

Biophysical Journal

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The lower critical solution temperature (LCST) of the thermo-responsive engineered elastin-like polypeptide (ELP) biopolymer is being exploited for the thermal targeted delivery of doxorubicin (Dox) to solid tumors. We examine the impact of Dox labeling on the thermodynamic and hydrodynamic behavior of an ELP drug carrier and how Dox influences the liquid-liquid phase separation (LLPS). Turbidity, dynamic light scattering (DLS), and differential scanning calorimetry measurements show that ELP undergoes a cooperative liquid-liquid phase separation from a soluble to insoluble coacervated state that is enhanced by Dox labeling. Circular dichroism measurements show that below the LCST ELP consists of both random coils and temperature-dependent b-turn structures. Labeling with Dox further enhances b-turn formation. DLS measurements reveal a significant increase in the hydrodynamic radius of ELP below the LCST consistent with weak self-association. Doxlabeled SynB1-ELP1 (Dox-ELP) has a significant increase in the hydrodynamic radius by DLS measurements that is consistent with stable oligomers and, at high Dox-ELP concentrations, micelle structures. Enhanced association by Dox-ELP is confirmed by sedimentation velocity analytical ultracentrifugation measurements. Both ELP self-association and the ELP inverse phase transition are entropically driven with positive changes in enthalpy and entropy. We show by turbidity and DLS that the ELP phase transition is monophasic, whereas mixtures of ELP and Dox-ELP are biphasic, with Dox-labeled ELP phase changing first and unlabeled ELP partitioning into the coacervate as the temperature is raised. DLS reveals a complex growth in droplet sizes consistent with coalescence and fusion of liquid droplets. Differential scanning calorimetry measurements show a À11 kcal/mol change in enthalpy for Dox-ELP coacervation relative to the unlabeled ELP, consistent with droplet formation being stabilized by favorable enthalpic interactions. We propose that the ELP phase change is initiated by ELP self-association, enhanced by increased Dox-ELP oligomer and micelle formation and stabilized by favorable enthalpic interactions in the liquid droplets.

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“…One manuscript addresses the phenomenon of liquidliquid phase separation by IDPs, a topic of growing interest in the past few years, as many IDPs have been observed drive the formation of biomolecular condensates both in vitro and in vivo. This study (7) used NMR to identify regions of secondary structure propensity in an IDP critical to intermolecular interactions in the early stages of phase separation.…”

mentioning

confidence: 99%

Proteins: Disorder, Folding, and Crowding

Rhoades

2019

Biophysical Journal

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The protein side of the central dogma of molecular biology holds that the native state of a protein is encoded by its sequence of amino acids. For more than 50 years, this principle has been largely applied to the folding of globular proteins. Both experimental and computational biophysicists have worked to determine the thermodynamic and kinetic parameters that yield underlying principles of protein folding, as well as to develop algorithms to successfully predict structure from sequence. More recently, the biophysical community has recognized that the native states of many proteins do not involve stable globular structures, but rather disordered, dynamic conformational ensembles. These intrinsically disordered proteins (IDPs) are involved in a broad range of cellular functions and assemblies, as well as having causal roles in a number of devastating human diseases. Disordered proteins and regions can be predicted by their amino sequences and even a single point mutation can disrupt their normal function. In other words, even when the native state of a protein does not involve stable tertiary contacts and canonical secondary structure, the central dogma still applies. Here, I highlight recent articles in the Biophysical Journal that represent advances and biophysical insights focusing on protein folding and IDPs.Coupled folding and binding of some IDPs upon interaction is at the intersection of the protein folding and IDP fields. This phenomenon is investigated in two recent papers in the journal. In the first (1), co-translational interactions between two IDPs of opposite charges are investigated by fluorescence correlation spectroscopy and force measurements. The second paper (2) seeks to tease apart the recognition and folding steps of an IDP for its heterogeneously structured partner, by systematic mutation of the IDP. While many small, globular proteins fold spontaneously in vitro, in the complex environment of the cell, proteins interact with a variety of biomolecules and many rely on a variety of chaperone molecules in order to fold properly.Two papers address protein folding in the presence of molecular chaperones. One of these (3) made circularly permuted variants of GFP which changed its contact order and characterized the interaction and folding of these variants with the chaperonin, GroEL. Fundamental aspects of surfactant mediated protein unfolding were studied in the second paper (4), where both ionic and hydrophobic properties of the surfactant were found to be important for unfolding an uncharged protein.Two papers address fundamental biophysical properties of IDPs. Mukhopadhyay and coworkers (5) used a combination of experiments and simulations to create a hydration map of a model IDP, finding dynamically distinct groups of water molecules associated with the protein. A computational study (6) looked at different types of molecular crowders on the conformational ensembles sampled by an IDP, observing both compact and extended conformations that interact differentially with the crowding macromole...

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Modeling the Early Stages of Phase Separation in Disordered Elastin-like Proteins

Cited by 18 publications

References 96 publications

Trends in the design and use of elastin-like recombinamers as biomaterials

Trends in the design and use of elastin-like recombinamers as biomaterials

Effects of Doxorubicin on the Liquid-Liquid Phase Change Properties of Elastin-Like Polypeptides

Proteins: Disorder, Folding, and Crowding

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