Dynamic metastable long-living droplets formed by sticker-spacer proteins

Multivalent biopolymers phase separate into membrane-less organelles (MLOs) which exhibit liquid-like behavior. Here, we explore formation of prototypical MOs from multivalent proteins on various time and length scales and show that the kinetically arrested metastable multi-droplet state is a dynamic outcome of the interplay between two competing processes: a diffusion-limited encounter between proteins, and the exhaustion of available valencies within smaller clusters. Clusters with satisfied valencies cannot coalesce readily, resulting in metastable, long-living droplets. In the regime of dense clusters akin to phase-separation, we observe co-existing assemblies, in contrast to the single, large equilibrium-like cluster. A system-spanning network encompassing all multivalent proteins was only observed at high concentrations and large interaction valencies. In the regime favoring large clusters, we observe a slow-down in the dynamics of the condensed phase, potentially resulting in loss of function. Therefore, metastability could be a hallmark of dynamic functional droplets formed by sticker-spacer proteins.

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Electron Incoherent Scattering from Nanometer-Sized Charge Ordering in Colossal Magnetoresistive La2/3ca1/3mnO3

Microscopy and Microanalysis ◽

10.1017/s1431927600028245 ◽

2001 ◽

Vol 7 (S2) ◽

pp. 434-435

Author(s):

J. M. Zuo

Keyword(s):

Phase Separation ◽

Electron Diffraction ◽

Degrees Of Freedom ◽

Complex Oxides ◽

Charge Ordering ◽

Phase Separations ◽

Length Scales ◽

Colossal Magnetoresistive ◽

Electronic Phase Separation ◽

Electronic Phase

Electronic phase separation is known to occur in complex oxides ranging from high-Tc superconductors to colossal magnetoresisitive (CMR) manganites. Accumulating experimental evidences show regions of temperature dependent conducting and insulating regions, whose exact origin is unknown. Theoretically, it is has been shown that these systems are unstable from the strong interplay between the lattice, charge and spin degrees of freedom.The key to understand the electronic phase separation in complex oxides is the structure. Electron diffraction is the only probe that covers the length scales from angstroms to microns. Characterization at these length scales is critical (electronic phase separations are typically about nanometers in sizes). Traditionally, electron diffraction has been played important roles in discovering the new types of phase separations, but has contributed little to the quantitative understanding. The reason is the strong interaction of electrons with matter, which gives both strong inelastic background and multiple scattering.

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How does organic structure determine organic reactivity? The effect of ortho-dimethyl groups on the nucleophilic substitution and alkene-forming elimination reactions of ring-substituted cumyl derivatives

Canadian Journal of Chemistry ◽

10.1139/v99-084 ◽

1999 ◽

Vol 77 (5-6) ◽

pp. 922-933 ◽

Cited By ~ 3

Author(s):

Tina L Amyes ◽

Tadeusz Mizerski ◽

John P Richard

Keyword(s):

Rate Constant ◽

Nucleophilic Addition ◽

Substituent Effects ◽

Fold Increase ◽

Methyl Groups ◽

Diffusion Limited ◽

Azide Ion ◽

Brønsted Base ◽

High Concentrations ◽

Ortho Substituent

The addition of a pair of ortho-methyl groups to ring-substituted cumyl derivatives to give the corresponding 2,6-dimethylcumyl derivatives X-1-Y leads to modest (<5-fold) changes in the observed rate constant for reaction in 50:50 (v:v) trifluoroethanol-water (I = 0.50, NaClO4). The reactions of X-1-Y proceed by a stepwise mechanism through the liberated 2,6-dimethylcumyl carbocations X-2 that partition between nucleophilic addition of solvent and deprotonation to give good yields ([Formula: see text] 67%) of the corresponding 2-(2,6-dimethylaryl)propenes X-3. The carbocations X-2 are also trapped by nucleophilic addition of azide ion to give good yields ([Formula: see text] 68% at [N3-] = 0.50 M) of the corresponding 2,6-dimethylcumyl azides X-1-N3. In the presence of high concentrations of azide ion there are constant limiting yields of the alkenes X-3, which shows that X-2 also undergo significant reactions with azide ion as a Brønsted base. The product rate constant ratios for partitioning of the 2,4,6-trimethylcumyl carbocation Me-2 between reaction with azide ion as a Lewis and a Brønsted base, kaz/kB, the nucleophilic addition of azide ion and solvent, kaz/ks (M-1), and deprotonation by solvent, kaz/ke (M-1), were combined with (kaz + kB) = 5 × 109 M-1 s-1 for the diffusion-limited reaction of azide ion to give absolute rate constants for the reactions of Me-2. The data show that the addition of a pair of ortho-methyl groups to the 4-methylcumyl carbocation to give the sterically hindered Me-2 results in a 70-fold decrease in the rate constant for nucleophilic addition of solvent to the benzylic carbon, but a 60-fold increase in the rate constant for deprotonation of the carbocation by solvent.Key words: carbocation, ortho-substituent effects, steric effects, solvolysis, elimination.

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Integrated diagnostic/prognostic tools for small cracks in structures

Proceedings of the Institution of Mechanical Engineers Part C Journal of Mechanical Engineering Science ◽

10.1243/0954406001523579 ◽

2000 ◽

Vol 214 (9) ◽

pp. 1123-1140 ◽

Cited By ~ 1

Author(s):

D. L. McDowell ◽

J. D. Clayton ◽

V. P. Bennett

Keyword(s):

Hot Spots ◽

High Cycle Fatigue ◽

Integrated System ◽

Loss Of Function ◽

Length Scales ◽

Critical Dimensions ◽

Material Defects ◽

Growth Laws ◽

Multiple Sensor ◽

Small Cracks

An integrated system for diagnosis of the ‘health’ of a structural component subjected to high-cycle fatigue (HCF) consists of sets of embedded or emplaced sensors at various locations, extracting information related to the generation of material defects, the presence of crack-like discontinuities and their progression and changes in system dynamics that may relate to this progression. Conceptually, signals from these sensors are fed into a processing environment that can project deleterious conditions related to the onset of loss of function or propagation of cracks to critical dimensions. Since the idea is to monitor the gradual changes of component performance and various local related indices before catastrophic failure to enable the operator to respond with a maintenance hold, it is essential to couple the diagnostics with prognostic capability; this facilitates a prediction of how much time remains within the window of viable servicing or repair. In the HCF regime, the dominant fraction of total fatigue life may be spent at crack lengths of the order of 20-500 um. The detection of longer cracks near the end of component life is critical since component failure may lead to failure of the overall structure. This necessitates the identification of (a) algorithms for identifying component ‘hot spots’ where failure is likely to occur, (b) development of appropriate crack growth laws for cracks of different length scales, ranging from the order of grain size to the order of component dimensions, including consideration of contacting components (fretting fatigue) and environmental effects and (c) development of algorithms for identifying the progression of component degradation on the basis of multiple-sensor inputs at different time and length scales, providing feedback to support cause for maintenance shutdown. This paper discusses each of these issues.

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Diverging length scales in diffusion-limited aggregation

Physical Review A ◽

10.1103/physreva.34.2558 ◽

1986 ◽

Vol 34 (3) ◽

pp. 2558-2560 ◽

Cited By ~ 55

Author(s):

Paul Meakin ◽

Fereydoon Family

Keyword(s):

Diffusion Limited Aggregation ◽

Length Scales ◽

Diffusion Limited

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Decreased piRNAs in Infertile Semen Are Related to Downregulation of Sperm MitoPLD Expression

Frontiers in Endocrinology ◽

10.3389/fendo.2021.696121 ◽

2021 ◽

Vol 12 ◽

Author(s):

Yeting Hong ◽

Yanqian Wu ◽

Jianbin Zhang ◽

Chong Yu ◽

Lu Shen ◽

...

Keyword(s):

Male Infertility ◽

Sperm Motility ◽

Seminal Plasma ◽

Molecular Mechanisms ◽

Molecular Biomarkers ◽

Loss Of Function ◽

Human Seminal Plasma ◽

High Concentrations ◽

Tight Correlation ◽

Infertile Patients

Currently, the molecular mechanisms underlining male infertility are still poorly understood. Our previous study has demonstrated that PIWI-interacting RNAs (piRNAs) are downregulated in seminal plasma of infertile patients and can serve as molecular biomarkers for male infertility. However, the source and mechanism for the dysregulation of piRNAs remain obscure. In this study, we found that exosomes are present in high concentrations in human seminal plasma and confirmed that piRNAs are predominantly present in the exosomal fraction of seminal plasma. Moreover, we showed that piRNAs were significantly decreased in exosomes of asthenozoospermia patients compared with normozoospermic men. By systematically screening piRNA profiles in sperms of normozoospermic men and asthenozoospermia patients, we found that piRNAs were parallelly reduced during infertility. At last, we investigated the expression of some proteins that are essential for piRNAs biogenesis in sperms and therefore identified a tight correlation between the levels of spermatozoa piRNA and MitoPLD protein, suggesting that the loss-of-function of MitoPLD could cause a severe defect of piRNA accumulation in sperms. In summary, this study identified a parallel reduction of piRNAs and MitoPLD protein in sperms of asthenozoospermia patients, which may provide pathophysiological clues about sperm motility.

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Three archetypical classes of macromolecular regulators of protein liquid–liquid phase separation

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.1907849116 ◽

2019 ◽

Vol 116 (39) ◽

pp. 19474-19483 ◽

Cited By ~ 22

Author(s):

Archishman Ghosh ◽

Konstantinos Mazarakos ◽

Huan-Xiang Zhou

Keyword(s):

Phase Separation ◽

Liquid Phase ◽

Computational Study ◽

Quantitative Measure ◽

Liquid Phase Separation ◽

Wide Range ◽

Regulatory Effects ◽

High Concentrations ◽

Droplet Phase ◽

Liquid Liquid Phase Separation

Membraneless organelles, corresponding to the droplet phase upon liquid–liquid phase separation (LLPS) of protein or protein–RNA mixtures, mediate myriad cellular functions. Cells use a variety of biochemical signals such as expression level and posttranslational modification to regulate droplet formation and dissolution, but the physical basis of the regulatory mechanisms remains ill-defined and quantitative assessment of the effects is largely lacking. Our computational study predicted that the strength of attraction by droplet-forming proteins dictates whether and how macromolecular regulators promote or suppress LLPS. We experimentally tested this prediction, using the pentamers of SH3 domains and proline-rich motifs (SH35 and PRM5) as droplet-forming proteins. Determination of the changes in phase boundary and the partition coefficients in the droplet phase over a wide range of regulator concentrations yielded both a quantitative measure and a mechanistic understanding of the regulatory effects. Three archetypical classes of regulatory effects were observed. Ficoll 70 at high concentrations indirectly promoted SH35–PRM5 LLPS, by taking up volume in the bulk phase and thereby displacing SH35 and PRM5 into the droplet phase. Lysozyme had a moderate partition coefficient and suppressed LLPS by substituting weaker attraction with SH35 for the stronger SH35–PRM5 attraction in the droplet phase. By forming even stronger attraction with PRM5, heparin at low concentrations partitioned heavily into the droplet phase and promoted LLPS. These characteristics were recapitulated by computational results of patchy particle models, validating the identification of the 3 classes of macromolecular regulators as volume-exclusion promotors, weak-attraction suppressors, and strong-attraction promotors.

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Pelle kinase is activated by autophosphorylation during Toll signaling in Drosophila

Development ◽

10.1242/dev.129.8.1925 ◽

2002 ◽

Vol 129 (8) ◽

pp. 1925-1933 ◽

Cited By ~ 1

Author(s):

Baohe Shen ◽

James L. Manley

Keyword(s):

Dependent Manner ◽

Loss Of Function ◽

Concentration Dependent Manner ◽

Downstream Target ◽

Toll Signaling ◽

Early Embryos ◽

High Concentrations ◽

L2 Cells

The Drosophila Pelle kinase plays a key role in the evolutionarily conserved Toll signaling pathway, but the mechanism responsible for its activation has been unknown. We present in vivo and in vitro evidence establishing an important role for concentration-dependent autophosphorylation in the signaling process. We first show that Pelle phosphorylation can be detected transiently in early embryos, concomitant with activation of signaling. Importantly, Pelle phosphorylation is enhanced in a gain-of-function Toll mutant (Toll10b), but decreased by loss-of-function Toll alleles. Next we found that Pelle is phosphorylated in transfected Schneider L2 cells in a concentration-dependent manner such that significant modification is observed only at high Pelle concentrations, which coincide with levels required for phosphorylation and activation of the downstream target, Dorsal. Pelle phosphorylation is also enhanced in L2 cells co-expressing Toll10b, and is dependent on Pelle kinase activity. In vitro kinase assays revealed that recombinant, autophosphorylated Pelle is far more active than unphosphorylated Pelle. Importantly, unphosphorylated Pelle becomes autophosphorylated, and activated, by incubation at high concentrations. We discuss these results in the context of Toll-like receptor mediated signaling in both flies and mammals.

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