A comprehensive BIAPSS platform for the physicochemical featurization of phase separating proteins

Abstract The liquid-liquid phase separation (LLPS) of biological macromolecules has emerged as a foundational mechanism underlying the formation of a myriad of membraneless organelles (MLOs), such as stress granules, transcription factor condensates, and chromatin compartments. A molecular grammar of sequences, which would enable a quantitative prediction and understanding of protein phase separation from first principles is currently missing. A major challenge in the field is the sparsity of bioinformatics data and the lack of computational, data-driven tools for biophysical and statistical analysis of proteins capable of phase separation. Here we present the utility of web applications framed within a novel open-source platform for BioInformatic Analysis of liquid-liquid Phase-Separating protein Sequences, https://biapss.chem.iastate.edu/. BIAPSS combines high-throughput interactive data analytics of physicochemical and evolutionary features with a comprehensive repository of bioinformatic data for on-the-fly research of the sequence-dependent properties of proteins with known LLPS behavior. To facilitate exploration of the services and provide the interpretation guideline, we present two attention-getting case studies of FUS and hnRNPDL. This should help the LLPS community uncover the nature of interactions driving the formation of membraneless organelles.

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Charge-Driven Condensation of RNA and Proteins Suggests Broad Role of Phase Separation in Cytoplasmic Environments

10.1101/2020.04.23.057901 ◽

2020 ◽

Author(s):

Bercem Dutagaci ◽

Grzegorz Nawrocki ◽

Joyce Goodluck ◽

Ali Akbar Ashkarran ◽

Charles G. Hoogstraten ◽

...

Keyword(s):

Phase Separation ◽

Liquid Phase ◽

Molecular Size ◽

Liquid Phase Separation ◽

Coarse Grained ◽

Biological Macromolecules ◽

Separation Processes ◽

Cellular Environment ◽

Liquid Liquid Phase Separation

ABSTRACTPhase separation processes are increasingly being recognized as important organizing mechanisms of biological macromolecules in cellular environments. Well established drivers of liquid-liquid phase separation are multi-valency and intrinsic disorder. Here, we show that globular macromolecules may condense simply based on electrostatic complementarity. More specifically, phase separation of mixtures between RNA and positively charged proteins is described from a combination of multiscale computer simulations with microscopy and spectroscopy experiments. Condensates retain liquid character and phase diagrams are mapped out as a function of molecular concentrations in experiment and as a function of molecular size and temperature via simulations. The results suggest a more general principle for phase separation that is based primarily on electrostatic complementarity without invoking polymer properties as in most previous studies. Simulation results furthermore suggest that such phase separation may occur widely in heterogenous cellular environment between nucleic acid and protein components.STATEMENT OF SIGNIFICANCELiquid-liquid phase separation has been recognized as a key mechanism for forming membrane-less organelles in cells. Commonly discussed mechanisms invoke a role of disordered peptides and specific multi-valent interactions. We report here phase separation of RNA and proteins based on a more universal principle of charge complementarity that does not require disorder or specific interactions. The findings are supported by coarse-grained simulations, theory, and experimental validation via microscopy and spectroscopy. The broad implication of this work is that condensate formation may be a universal phenomenon in biological systems.

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BIAPSS - BioInformatic Analysis of liquid-liquid Phase-Separating protein Sequences

10.1101/2021.02.11.430806 ◽

2021 ◽

Author(s):

Aleksandra E. Badaczewska-Dawid ◽

Davit A. Potoyan

Keyword(s):

Phase Separation ◽

Liquid Phase ◽

Open Source ◽

Protein Sequences ◽

Bioinformatic Analysis ◽

Liquid Phase Separation ◽

Intrinsically Disordered Regions ◽

Web Platform ◽

Liquid Liquid Phase Separation ◽

In Cells

AbstractLiquid-liquid phase separation (LLPS) has recently emerged as a cornerstone mechanism underlying the biogenesis of membraneless organelles (MLOs). However, a quantitative molecular grammar of protein sequences that controls the LLPS remains poorly understood. The progress in this field is hampered by the insufficiency of comprehensive databases and associated computational infrastructure for targeting biophysical and statistical analysis of phase separating biopolymers. Therefore, we have created a novel open-source web platform named BIAPSS (BioInformatic Analysis of liquid-liquid Phase-Separating protein Sequences) which contains interactive data analytic tools in combination with a comprehensive repository of bioinformatic data for on-the-fly exploration of sequence-dependent properties of proteins with known LLPS behavior. BIAPSS includes a residue-resolution biophysical analyzer for interrogating individual protein sequences (SingleSEQ tab). The latter allows users to correlate regions prone to phase separation with a large array of physicochemical attributes and various short linear motifs. BIAPSS also includes global statistics derived over the universe of most of the known LLPS-driver protein sequences (MultiSEQ tab) for revealing the regularities and sequence-specific signals driving phase separation. Finally, BIAPSS incorporates an extensive cross-reference section that links all entries to primary LLPS databases and other external resources thereby serving as a central navigation hub for the phase separation community. All of the data used by BIAPSS is freely available for download as well-formatted pre-processed data with detailed descriptions, facilitating rapid implementation in user-defined computational protocols.Abstract FigureTOC - graphical abstractAuthor summaryProteins, especially those with low complexity and intrinsically disordered regions, have recently come into the limelight because of mounting evidence showing that these regions can drive the formation of membraneless organelles (MLOs) in cells. The underlying physical mechanism for forming MLOs is liquid-liquid phase separation (LLPS); a thermodynamically driven process whereby a cellular milieu with a relatively well-mixed distribution of biomolecules gets decomposed into liquid droplets where the concentration of selected biomolecules is higher. Deciphering molecular sequence grammar of phase separation has turned out to be challenging because of the complexity of this process in cells and the vastness of sequence space of LLPS-driver proteins. While the field is still in its infancy the growth of experimental data has already spurred the creation of several major databases which collect and annotate bimolecular systems with confirmed LLPS behavior. What is currently missing is a framework that would leverage the existing databases by integrating them with deep biophysical and bioinformatic analysis for identifying statistically significant features of protein sequences implicated in LLPS. In this work, we have addressed this challenge by creating an open-source web platform named BIAPSS (BioInformatic Analysis of liquid-liquid Phase-Separating protein Sequences) which integrates a comprehensive repository of pre-processed bioinformatic data for LLPS-driver protein sequences with interactive analytic applications for on-the-fly analysis of biophysical features relevant for LLPS behavior. BIAPSS empowers users with novel and effective tools for exploring LLPS-related sequence signals for individual proteins (SingleSEQ tab) and globally by integrating common regularities across subgroups or the entire LLPS sequence superset (MultiSEQ). The long-term plan for BIAPSS is to serve as a unifying hub for the experimental and computational community with a comprehensive set of analytic tools, biophysically featured data, and standardized protocols facilitating the identification of sequence hot spots driving the LLPS, which all can support applications for designing new sequences of biomedical interest.

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Relevance of Liquid-Liquid Phase Separation of Supersaturated Solution in Oral Absorption of Albendazole from Amorphous Solid Dispersions

Pharmaceutics ◽

10.3390/pharmaceutics13020220 ◽

2021 ◽

Vol 13 (2) ◽

pp. 220

Author(s):

Kyosuke Suzuki ◽

Kohsaku Kawakami ◽

Masafumi Fukiage ◽

Michinori Oikawa ◽

Yohei Nishida ◽

...

Keyword(s):

Phase Separation ◽

Liquid Phase ◽

Oral Absorption ◽

Hydroxypropyl Methylcellulose ◽

Amorphous Solid ◽

Liquid Phase Separation ◽

Quantitative Prediction ◽

Amorphous Solid Dispersion ◽

Supersaturated Solution ◽

Liquid Liquid Phase Separation

Amorphous solid dispersion (ASD) is one of the most promising formulation technologies for improving the oral absorption of poorly soluble drugs, where the maintenance of supersaturation plays a key role in enhancing the absorption process. However, quantitative prediction of oral absorption from ASDs is still difficult. Supersaturated solutions can cause liquid-liquid phase separation through the spinodal decomposition mechanism, which must be adequately comprehended to understand the oral absorption of drugs quantitatively. In this study, albendazole (ALZ) was formulated into ASDs using three types of polymers, poly(methacrylic acid-co-methyl methacrylate) (Eudragit) L100, Vinylpyrrolidone-vinyl acetate copolymer (PVPVA), and hydroxypropyl methylcellulose acetate succinate (HPMCAS). The oral absorption of ALZ in rats administered as ASD suspensions was not explained by dissolution study but was predicted using liquid-liquid phase separation concentration, which suggested that the absorption of ALZ was solubility-limited. The oral administration study in dogs performed using solid capsules demonstrated the low efficacy of ASDs because the absorption was likely to be limited by dissolution rate, which indicated the importance of designing the final dosage form of the ASDs.

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Driving Forces of Liquid–Liquid Phase Separation in Biological Systems

Biomolecules ◽

10.3390/biom9090473 ◽

2019 ◽

Vol 9 (9) ◽

pp. 473

Author(s):

Zaslavsky ◽

Ferreira ◽

Uversky

Keyword(s):

Phase Separation ◽

Liquid Phase ◽

Interfacial Tension ◽

Driving Forces ◽

Biological Systems ◽

Liquid Phase Separation ◽

Necessary Condition ◽

Biological Macromolecules ◽

Solvent Properties ◽

Liquid Liquid Phase Separation

Analysis of liquid–liquid phase separation in biological systems shows that this process is similar to the phase separation observed in aqueous two-phase systems formed by nonionic polymers, proteins, and polysaccharides. The emergence of interfacial tension is a necessary condition of phase separation. The situation in this regard is similar to that of phase separation in mixtures of partially miscible solvents. It is suggested that the evaluation of the effects of biological macromolecules on the solvent properties of aqueous media and the measurement of the interfacial tension as a function of these solvent properties may be more productive for gaining insights into the mechanism of liquid–liquid phase separation than the study of structural details of proteins and RNAs engaged in the process.

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Faculty Opinions recommendation of A lipid bound actin meshwork organizes liquid phase separation in model membranes.

Faculty Opinions – Post-Publication Peer Review of the Biomedical Literature ◽

10.3410/f.718314286.793492585 ◽

2014 ◽

Author(s):

Robert Piper

Keyword(s):

Phase Separation ◽

Liquid Phase ◽

Liquid Phase Separation ◽

Model Membranes

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Faculty Opinions recommendation of Mouse Heterochromatin Adopts Digital Compaction States without Showing Hallmarks of HP1-Driven Liquid-Liquid Phase Separation.

Faculty Opinions – Post-Publication Peer Review of the Biomedical Literature ◽

10.3410/f.737432801.793572772 ◽

2020 ◽

Author(s):

Sophie Polo ◽

Pierre Caron

Keyword(s):

Phase Separation ◽

Liquid Phase ◽

Liquid Phase Separation ◽

Liquid Liquid Phase Separation

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Recent advances in hyphenation of nanoflow liquid phase separation with inductively coupled plasma mass spectrometry

Chinese Journal of Chromatography ◽

10.3724/sp.j.1123.2018.11025 ◽

2019 ◽

Vol 37 (2) ◽

pp. 132

Author(s):

Fangfang DING ◽

Jue ZHU ◽

Rui GUO ◽

Bo ZHANG

Keyword(s):

Mass Spectrometry ◽

Phase Separation ◽

Liquid Phase ◽

Inductively Coupled Plasma ◽

Liquid Phase Separation ◽

Inductively Coupled ◽

Recent Advances ◽

Plasma Mass Spectrometry

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Focusing on mRNA Granules and Stalled Polysomes Amidst Diverse Mechanisms Underlying mRNA Transport, mRNA Storage, and Local Translation

The Oxford Handbook of Neuronal Protein Synthesis ◽

10.1093/oxfordhb/9780190686307.013.20 ◽

2020 ◽

Author(s):

Mina N. Anadolu ◽

Wayne S. Sossin

Keyword(s):

Protein Synthesis ◽

Liquid Phase ◽

Mrna Translation ◽

Rna Transport ◽

P Bodies ◽

Rna Granules ◽

Mrna Granules ◽

Transport Particle ◽

As Stress ◽

The Individual

In neurons, mRNAs are transported to distal sites to allow for localized protein synthesis. There are many diverse mechanisms underlying this transport. For example, an individual mRNA can be transported in an RNA transport particle that is tailored to the individual mRNA and its associated binding proteins. In contrast, some mRNAs are transported in liquid-liquid phase separated structures called neuronal RNA granules that are made up of multiple stalled polysomes, allowing for rapid initiation-independent production of proteins required for synaptic plasticity. Moreover, neurons have additional types of liquid-liquid phase–separated structures containing mRNA, such as stress granules and P bodies. This chapter discusses the relationships between all of these structures, what proteins distinguish them, and the possible roles they play in the complex control of mRNA translation at distal sites that allow neurons to use protein synthesis to refine their local proteome in many different ways.

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