Multivalent Interactions between Molecular Components Involved in Clathrin Independent Endocytosis Drive Protein Phase Separation

Endocytosis of transmembrane receptors initiates via molecular interactions between the activated receptor and the endocytic machinery. A specific group of receptors, including the β1-adrenergic receptor (β1-AR), is internalized through a non-clathrin pathway known as Fast Endophilin Mediated Endocytosis (FEME). A key question is: how does the endocytic machinery assemble and how is it modulated by activated receptors during FEME. Here we show that endophilin, a major regulator of FEME, undergoes a phase transition into liquid-like condensates, which facilitates the formation of multi-protein assemblies by enabling the phase partitioning of endophilin binding proteins. The phase transition can be triggered by specific multivalent binding partners of endophilin in the FEME pathway such as the third intracellular loop (TIL) of the β1-AR, and the proline-rich-motifs of lamellipodin (LPD-PRMs). Other endocytic accessory proteins can either partition into, or target interfacial regions of, these condensate droplets. On the membrane, TIL promotes protein clustering in the presence of endophilin and LPD-PRMs. Our results demonstrate how the multivalent interactions between endophilin, LPD-PRMs and TIL regulate protein assembly formation on the membrane, providing mechanistic insights into the priming and initiation steps of FEME.

Strong deviations from thermodynamically expected phase partitioning of organic acids during one year of rural field measurements

<p>Organic acids are ubiquitous compounds in the troposphere and can affect human health, the climate, air quality, and the linked ecosystems. Depending on their solubility and volatility, they can partition in both gas phase and in the particle phase. In the particle phase, organic acids partly represent about 10% of the water-soluble organic matter. However, their partitioning between different phases is not fully understood yet. Therefore, an upgraded monitor for aerosols and gases in ambient air (MARGA) was applied for one year at the Central European TROPOS research site Melpitz to study the gas- and particle-phase partitioning of formic, acetic, propionic, butyric, glycolic, pyruvic, oxalic, malonic, succinic, malic, and methanesulfonic acid (MSA). Measured gas- and PM<sub>10</sub> particle-phase mean concentrations were 12−445 and 7−31 ng m<sup>-3</sup> for monocarboxylic acids (MCAs), between 0.6−8 and 4−31 ng m<sup>-3</sup> for dicarboxylic acids (DCAs), and 2 and 31 ng m<sup>-3</sup> for MSA, respectively. Assuming full dissolution in nonideal aerosol solutions, empirical noneffective Henry’s law constants (H<sub>emp</sub>) were calculated and compared with literature values (H<sub>lit</sub>). Calculated mean H<sub>emp</sub> were 4.5 × 10<sup>9</sup>−2.2 × 10<sup>10</sup> mol L<sup>−1</sup> atm<sup>−1</sup> for MCAs, 3.6 × 10<sup>10</sup>−7.5 × 10<sup>11</sup> mol L<sup>−1</sup> atm<sup>−1</sup> for DCAs, and 7.5 × 10<sup>7</sup> mol L<sup>−1</sup> atm<sup>−1</sup> for MSA and, thus, factors of 5.1 × 10<sup>3</sup>−9.1 × 10<sup>5</sup> and 2.5−20.3 higher than their corresponding H<sub>lit</sub> for MCAs and DCAs, respectively, and 9.0 × 10<sup>−5</sup> lower than H<sub>lit,MSA</sub>. Data analyses and thermodynamic calculations implicate that the formation of chemical association complexes and organic salts inhibits the partitioning of organic acids toward the gas phase and, thus, at least partly explains higher H<sub>emp</sub> values for both MCAs and summertime DCAs. Low H<sub>emp,MSA</sub> are also unexpected because of the high MSA solubility and are reported for the first time in this study. Overall, the results of the present study implicate that processes responsible for the observed stronger partitioning of carboxylic acids toward the particle phase need to be further investigated and accounted for in complex multiphase chemistry models as they affect the contribution of organic acids to secondary organic aerosol mass, their chemical processing, and lifetime.</p> <p> </p> <p> </p>

Weak Multivalent Biomolecular Interactions: A Strength vs Numbers Tug of War with Implications for Phase Partitioning

In this short perspective, we discuss how recent dynamic live cell imaging experiments have challenged our understanding of mechanisms driving functional molecular interactions in vivo. While we have generally considered the formation of functional biomolecular complexes as resulting from the stable assembly of two or more partner molecules, here we entertain the possibility that function may actually be maintained while molecules are rapidly exchanged within a complex. We postulate that at high effective concentrations, even very weak interactions can lead to strong binding site occupancy and thereby, mediate function in a highly dynamic fashion. This new perspective in our definition of what represents a functional complex in living cells and in vivo could significantly alter how we define the nature of molecular transactions critical for mediating regulation in the cellular context. These less conventional principles also allow a broadening of the mechanistic options we should explore when interpreting essential biological processes such as gene regulation.

Latest Advances in Protein-Recovery Technologies from Agricultural Waste

In recent years, downstream bioprocessing industries are venturing into less tedious, simple, and high-efficiency separation by implementing advanced purification and extraction methods. This review discusses the separation of proteins, with the main focus on amylase as an enzyme from agricultural waste using conventional and advanced techniques of extraction and purification via a liquid biphasic system (LBS). In comparison to other methods, such as membrane extraction, precipitation, ultrasonication, and chromatography, the LBS stands out as an efficient, cost-effective, and adaptable developing method for protein recovery. The two-phase separation method can be water-soluble polymers, or polymer and salt, or alcohol and salt, which is a simpler and lower-cost method that can be used at a larger purification scale. The comparison of different approaches in LBS for amylase purification from agricultural waste is also included. Current technology has evolved from a simple LBS into microwave-assisted LBS, liquid biphasic flotation (LBF), thermoseparation (TMP), three-phase partitioning (TPP), ultrasound-assisted LBS, and electrically assisted LBS. pH, time, temperature, and concentration are some of the significant research parameters considered in the review of advanced techniques.

Solid-Phase Partitioning and Leaching Behavior of Pb and Zn from Playground Soils in Kabwe, Zambia

Zambia's Kabwe mine wastes (KMWs) are responsible for contaminating the surrounding soil and dust in the Kabwe district. Unfortunately, these wastes arise from the historical mining activities of lead (Pb) and Zinc (Zn), which lacked adequate waste management strategies. As a result, potentially toxic elements (PTEs) (Pb and Zn) spread across the Kabwe district. To assess the soil pollution derived from previous mining activities, we studied topsoil samples (n = 8) from the school playground soils (SPs). In this study, the degree of contamination, geochemical partitioning, and leachability, coupled with the release and retention of Pb and Zn, were studied. The SPs were classified as extremely enriched (EF > 40) and contaminated with Pb (Igeo > 5). On average, Pb (up to 89%) and Zn (up to 69%) were bound with exchangeable, weak acid-soluble, reducible and oxidizable phases, which are considered as 'geochemically mobile' phases in the environment. The leachates from the soils (n = 5) exceeded the Zambian standard (ZS: 190:2010) for Pb in potable drinking water (Pb < 0.01 mg/L). Furthermore, the spatial distribution of Pb and Zn showed a significant reduction in contents of Pb and Zn with the distance from the mine area.