Dynamic behavior of liquid droplets with enzyme compartmentalization triggered by sequential glycolytic enzyme reactions.
23 Nov 2021-Chemical Communications (The Royal Society of Chemistry)-Vol. 57, Iss: 93, pp 12544-12547
TL;DR: In this article, a model LLPS system coupled with a sequential glycolytic enzymatic reaction was developed to reproduce the dynamic control of liquid droplets; the droplets, which consist of poly-L-lysine and nucleotides, compartmentalize two different enzymes (hexokinase and glucose-6-phosphate dehydrogenase) individually, accelerating the overall reaction.
About: This article is published in Chemical Communications.The article was published on 2021-11-23. It has received 6 citations till now.
Citations
More filters
TL;DR: In this article , a wide variety of small anionic metabolites can form complex coacervate protocells with oligoarginine (R10) by phase separation, and they reveal the intricate interplay between prebiotically relevant molecules that combine these elements can provide insight into the requirements for the formation of life-like protocells from abiotic building blocks.
Abstract: Metabolism and compartmentalization are two of life's most central elements. Constructing synthetic assemblies based on prebiotically relevant molecules that combine these elements can provide insight into the requirements for the formation of life-like protocells from abiotic building blocks. In this work, we show that a wide variety of small anionic metabolites can form complex coacervate protocells with oligoarginine (R10) by phase separation. The coacervate stability can be rationalized by the molecular structure of the metabolites, and we show that three negative charges for carboxylates, or two negative charges complemented with an unsaturated moiety for phosphates and sulfates is sufficient for phase separation. The metabolites remain reactive after compartmentalization, and we show that protometabolic reactions can induce coacervate formation. The resulting coacervates can localize other metabolites and enhance their conversion. Finally, reactions of compartmentalized metabolites can also alter the physicochemical properties of the coacervates and ultimately lead to protocell dissolution. These results reveal the intricate interplay between (proto)metabolic reactions and coacervate compartments, and show that coacervates are excellent candidates for metabolically active protocells.
2 citations
TL;DR: The approach to quantify the contribution of non-ionic amino acids can be expected to help to provide a more accurate description and prediction of the LLPS propensity of peptides/proteins.
2 citations
18 Apr 2023
TL;DR: In this article , target-specific peptides are used to control liquid-liquid phase separation (LLPS) of proteins and modulate the physical nature of condensate, which can be used to gain deeper insights in the LLPS-mediated amyloid formation.
Abstract: Abstract Liquid-liquid phase separation (LLPS) of protein that leads to formation of membrane-less organelles is a critical event to many processes in the cell. Recently, some disease-related proteins, such as α-synuclein (αSyn), were found to undergo LLPS before their formation of amyloid fibrils. However, the progress towards controlling LLPS has been limited, and there has been no emerging engineered de novo molecules to induce and modulate the LLPS of targeted proteins. Here we report peptides that efficiently induce the LLPS of αSyn, discovered by the RaPID (random non-standard peptides integrated discovery) system. These peptides are able to co-localize with αSyn in liquid droplets via heterotypic interacting with the N- and C-terminal regions of αSyn. Our study demonstrates the capacity of target-specific peptides to control LLPS and modulate the physical nature of condensate. Thus, these peptides could be a unique tool to gain deeper insights in the LLPS-mediated amyloid formation.
TL;DR: In this article , the concentration of a G-quadruplex-forming RNA and an RGG peptide in a single droplet using Raman microscopy was found to maintain even when the introduced concentration ratio of these two species was varied.
Abstract: Liquid–liquid phase separation (LLPS), which forms highly concentrated droplets of biomolecules, is involved in various physiological phenomena. We have performed a label-free quantification of the concentration of a G-quadruplex-forming RNA and an RGG peptide in a single droplet using Raman microscopy. The concentration ratio of the RNA to the peptide within the droplet was found to maintain even when the introduced concentration ratio of these two species was varied. This result indicates that electrostatic interactions between the RNA and the peptide induced the droplet formation. It was also shown that the RNA maintains its G-quadruplex structure inside the droplets.
TL;DR: In this article , the Hsp90 and Hsp70 chaperone families act on human metabolic enzymes and their supramolecular assemblies to change enzymatic activities and metabolite flux.
References
More filters
TL;DR: This work has shown that liquid–liquid phase separation driven by multivalent macromolecular interactions is an important organizing principle for biomolecular condensates and has proposed a physical framework for this organizing principle.
Abstract: In addition to membrane-bound organelles, eukaryotic cells feature various membraneless compartments, including the centrosome, the nucleolus and various granules. Many of these compartments form through liquid–liquid phase separation, and the principles, mechanisms and regulation of their assembly as well as their cellular functions are now beginning to emerge. Biomolecular condensates are micron-scale compartments in eukaryotic cells that lack surrounding membranes but function to concentrate proteins and nucleic acids. These condensates are involved in diverse processes, including RNA metabolism, ribosome biogenesis, the DNA damage response and signal transduction. Recent studies have shown that liquid–liquid phase separation driven by multivalent macromolecular interactions is an important organizing principle for biomolecular condensates. With this physical framework, it is now possible to explain how the assembly, composition, physical properties and biochemical and cellular functions of these important structures are regulated.
3,294 citations
TL;DR: The findings together suggest that several membrane-less organelles have been shown to exhibit a concentration threshold for assembly, a hallmark of phase separation, and represent liquid-phase condensates, which form via a biologically regulated (liquid-liquid) phase separation process.
Abstract: BACKGROUND Living cells contain distinct subcompartments to facilitate spatiotemporal regulation of biological reactions. In addition to canonical membrane-bound organelles such as secretory vesicles and endoplasmic reticulum, there are many organelles that do not have an enclosing membrane yet remain coherent structures that can compartmentalize and concentrate specific sets of molecules. Examples include assemblies in the nucleus such as the nucleolus, Cajal bodies, and nuclear speckles and also cytoplasmic structures such as stress granules, P-bodies, and germ granules. These structures play diverse roles in various biological processes and are also increasingly implicated in protein aggregation diseases. ADVANCES A number of studies have shown that membrane-less assemblies exhibit remarkable liquid-like features. As with conventional liquids, they typically adopt round morphologies and coalesce into a single droplet upon contact with one another and also wet intracellular surfaces such as the nuclear envelope. Moreover, component molecules exhibit dynamic exchange with the surrounding nucleoplasm and cytoplasm. These findings together suggest that these structures represent liquid-phase condensates, which form via a biologically regulated (liquid-liquid) phase separation process. Liquid phase condensation increasingly appears to be a fundamental mechanism for organizing intracellular space. Consistent with this concept, several membrane-less organelles have been shown to exhibit a concentration threshold for assembly, a hallmark of phase separation. At the molecular level, weak, transient interactions between molecules with multivalent domains or intrinsically disordered regions (IDRs) are a driving force for phase separation. In cells, condensation of liquid-phase assemblies can be regulated by active processes, including transcription and various posttranslational modifications. The simplest physical picture of a homogeneous liquid phase is often not enough to capture the full complexity of intracellular condensates, which frequently exhibit heterogeneous multilayered structures with partially solid-like characters. However, recent studies have shown that multiple distinct liquid phases can coexist and give rise to richly structured droplet architectures determined by the relative liquid surface tensions. Moreover, solid-like phases can emerge from metastable liquid condensates via multiple routes of potentially both kinetic and thermodynamic origins, which has important implications for the role of intracellular liquids in protein aggregation pathologies. OUTLOOK The list of intracellular assemblies driven by liquid phase condensation is growing rapidly, but our understanding of their sequence-encoded biological function and dysfunction lags behind. Moreover, unlike equilibrium phases of nonliving matter, living cells are far from equilibrium, with intracellular condensates subject to various posttranslational regulation and other adenosine triphosphate–dependent biological activity. Efforts using in vitro reconstitution, combined with traditional cell biology approaches and quantitative biophysical tools, are required to elucidate how such nonequilibrium features of living cells control intracellular phase behavior. The functional consequences of forming liquid condensates are likely multifaceted and may include facilitated reaction, sequestration of specific factors, and organization of associated intracellular structures. Liquid phase condensation is particularly interesting in the nucleus, given the growing interest in the impact of nuclear phase behavior on the flow of genetic information; nuclear condensates range from micrometer-sized bodies such as the nucleolus to submicrometer structures such as transcriptional assemblies, all of which directly interact with and regulate the genome. Deepening our understanding of these intracellular states of matter not only will shed light on the basic biology of cellular organization but also may enable therapeutic intervention in protein aggregation disease by targeting intracellular phase behavior.
2,432 citations
TL;DR: In this paper, a phase separation model was proposed to explain established and recently described features of transcriptional control, such as the formation of super-enhancers, the sensitivity of superenhancers to perturbation, the transcriptional bursting patterns of enhancers, and the ability of an enhancer to produce simultaneous activation at multiple genes.
1,162 citations
TL;DR: It is demonstrated that protein phase separation can create a distinct physical and biochemical compartment that facilitates signaling and promote signaling outputs both in vitro and in human Jurkat T cells.
Abstract: Activation of various cell surface receptors triggers the reorganization of downstream signaling molecules into micrometer- or submicrometer-sized clusters. However, the functional consequences of such clustering have been unclear. We biochemically reconstituted a 12-component signaling pathway on model membranes, beginning with T cell receptor (TCR) activation and ending with actin assembly. When TCR phosphorylation was triggered, downstream signaling proteins spontaneously separated into liquid-like clusters that promoted signaling outputs both in vitro and in human Jurkat T cells. Reconstituted clusters were enriched in kinases but excluded phosphatases and enhanced actin filament assembly by recruiting and organizing actin regulators. These results demonstrate that protein phase separation can create a distinct physical and biochemical compartment that facilitates signaling.
853 citations
TL;DR: What is known about the coupling of translation and mRNA degradation, the properties of P-bodies and stress granules, and how assembly of mRNPs into larger structures might influence cellular function are reviewed.
Abstract: The control of translation and mRNA degradation is important in the regulation of eukaryotic gene expression. In general, translation and steps in the major pathway of mRNA decay are in competition with each other. mRNAs that are not engaged in translation can aggregate into cytoplasmic mRNP granules referred to as processing bodies (P-bodies) and stress granules, which are related to mRNP particles that control translation in early development and neurons. Analyses of P-bodies and stress granules suggest a dynamic process, referred to as the mRNA Cycle, wherein mRNPs can move between polysomes, P-bodies and stress granules although the functional roles of mRNP assembly into higher order structures remain poorly understood. In this article, we review what is known about the coupling of translation and mRNA degradation, the properties of P-bodies and stress granules, and how assembly of mRNPs into larger structures might influence cellular function.
642 citations