Biology subjects

Michas, A.

Publications and source records attributed to Michas, A..

5 recordsLinked to original sources

bioRxiv2026

GW182 silencing domain engages the tristetraprolin-binding pocket of CNOT1 to recruit CNOT1 into multiprotein condensates

GW182 recruits the CCR4-NOT deadenylase complex through its CIM1 region, yet the CNOT1 surface mediating this interaction has remained unknown. Using hydrogen- deuterium exchange mass spectrometry, we map CIM1 binding to a hydrophobic groove within CNOT1 residues 800-999. Unexpectedly, this groove coincides with the tristetraprolin (TTP)-binding pocket. CIM1 and the C-terminal TTP peptide engage this surface through a shared RL(P/{zeta})X{Omega} sequence pattern despite lacking overall homology, suggesting a convergently evolved short linear motif (SLiM). We further show that the GW182 silencing domain (GW182 SD) and TTP compete for CNOT1 binding using orthogonal in vitro assays, including a reconstituted liquid-liquid phase separation system in which GW182 SD acts as a scaffold and recruits CNOT1 as a client through specific binding. These findings define a shared CNOT1 recognition site and reveal SLiM-mediated competition as a molecular principle governing CCR4-NOT engagement, with consequences for GW182-driven silencing condensate assembly in vitro. SignificanceGW182 silencing domain binds the tristetraprolin pocket of CNOT1 and recruits it into multiprotein condensates, providing a biophysical framework for coordinated miRNA- and ARE-mediated gene expression regulation.

biophysics↗

bioRxiv2025

Electrostatic Collapse of Intrinsically Disordered Acid-Rich Protein Is Sensitive to Counterion Valency

Intrinsically disordered proteins (IDPs) respond sensitively to their ionic environment, yet the mechanisms driving ion-induced conformational changes remain incompletely understood. Here, we investigate how counterion valency modulates the dimensions of an extremely charged model IDP, the aspartic and glutamic acid-rich protein AGARP. Fluorescence correlation spectroscopy and size exclusion chromatography reveal a pronounced, valency-dependent reduction in its hydrodynamic radius, with divalent cations (Ca2+, Mg2+) inducing collapse at much lower activities than monovalent cations (Na+, K+). Molecular dynamics simulations, direct sampling, and polyampholyte theory quantitatively capture the Debye-Huckel screening by monovalent ions but not the enhanced compaction driven by divalent ion binding. Circular dichroism spectroscopy shows that compaction occurs without secondary structure formation. Our results demonstrate a structure-free electrostatic collapse and suggest that specific chelation of divalent ions by disordered polyanionic protein chains is a key mechanism regulating IDP compaction, with implications for understanding their behavior in biologically relevant ionic environments. TOC GRAPHIC O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=108 SRC="FIGDIR/small/663830v1_ufig1.gif" ALT="Figure 1"> View larger version (43K): org.highwire.dtl.DTLVardef@84fba8org.highwire.dtl.DTLVardef@a5f3aforg.highwire.dtl.DTLVardef@12194a7org.highwire.dtl.DTLVardef@1a68366_HPS_FORMAT_FIGEXP M_FIG C_FIG

biophysics↗

bioRxiv2024

Extremely charged coral protein AGARP regulates calcium carbonate growth through liquid phase separation

Biomineralization via the non-classical crystallization pathway is postulated to involve a transient liquid phase of calcium carbonate formed in the presence of polymers. In the context of coral biocalcification, these polymers may include coral acid-rich proteins (CARPs) secreted into the skeletal organic matrix. However, direct evidence for the existence of this liquid phase containing proteins is lacking. Here, we show that the intrinsically disordered aspartic and glutamic acid-rich protein (AGARP), the first CARP cloned from the Great Barrier Reef scleractinian coral Acropora millepora, can significantly influence early stages of CaCO3 nucleation and crystal growth through liquid-liquid phase separation. We introduce the concept of a biologically relevant crystallization precursor: a liquid protein-calcium condensate composed of CARP molecules and Ca2+ ions, which forms as a result of liquid-liquid phase separation in a crowded environment. Our work bridges the gap between the liquid phase separation and biomineralization research.

biophysics↗

bioRxiv2024

Hydrodynamic Radii of Intrinsically Disordered Proteins: Fast Prediction by Minimum Dissipation Approximation and Experimental Validation

The diffusion coefficients of globular and fully unfolded proteins can be predicted with high accuracy solely from their mass or chain length. However, this approach fails for intrinsically disordered proteins (IDPs) containing structural domains. We propose a rapid predictive methodology for estimating the diffusion coefficients of IDPs. The methodology uses accelerated conformational sampling based on self-avoiding random walks and includes hydrodynamic interactions between coarse-grained protein subunits, modeled using the generalized Rotne-Prager-Yamakawa approximation. To estimate the hydrodynamic radius, we rely on the minimum dissipation approximation recently introduced by Cichocki et al. Using a large set of experimentally measured hydrodynamic radii of IDPs over a wide range of chain lengths and domain contributions, we demonstrate that our predictions are more accurate than the Kirkwood approximation and phenomenological approaches. Our technique may prove valuable in predicting the hydrodynamic properties of both fully unstructured and multidomain disordered proteins. TOC Graphic O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=199 SRC="FIGDIR/small/578612v1_ufig1.gif" ALT="Figure 1"> View larger version (34K): org.highwire.dtl.DTLVardef@1e2943aorg.highwire.dtl.DTLVardef@7193c8org.highwire.dtl.DTLVardef@9d5bceorg.highwire.dtl.DTLVardef@f64f72_HPS_FORMAT_FIGEXP M_FIG C_FIG

biophysics↗

bioRxiv2023

Diversity of hydrodynamic radii of intrinsically disordered proteins

Intrinsically disordered proteins (IDPs) form an important class of biomolecules regulating biological processes in higher organisms. The lack of a fixed spatial structure facilitates them to perform their regulatory functions. Due to the possibility of large conformational changes of IDPs, the cellular milieu can also control productivity of biochemical reactions. From the biophysical point of view, IDPs are biopolymers with a broad configuration state space. The conformation of such a biopolymer depends on non-covalent interactions of its amino acid side chain groups at given temperature and chemical conditions. Thus, the hydrodynamic radius (Rh) of an IDP of a given polymer length (N) is a sequence- and environment-dependent variable. We have reviewed the literature values of hydrodynamic radii of IDPs determined experimentally by SEC, AUC, PFG NMR, DLS, and FCS, and complement them with our FCS results obtained for a series of protein fragments involved in regulation of human gene expression. The data collected herein show that the values of hydrodynamic radii of intrinsically disordered proteins can span the full space between the folded globular and denatured proteins in the Rh(N) diagram.

biophysics↗