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The partition behavior of single and double-point mutants of bacteriophage T4 lysozyme (T4 lysozyme) and staphylococcal nuclease A was examined in different aqueous two-phase systems (ATPSs) and studied by Solvent Interaction Analysis (SIA). Additionally, the solvent accessible surface area (SASA) of modeled mutants of both proteins was calculated. The in silico calculations and the in vitro analyses of the staphylococcal nuclease and T4 lysozyme mutants correlate, indicating that the partition analysis in ATPSs provides a valid descriptor (SIA signature) covering various protein features, such as structure, structural dynamics, and conformational stability. [ABSTRACT FROM AUTHOR]

The partition coefficients of seven low molecular weight compounds were tested in different aqueous two-phase systems. The ionic composition of each system included specific salt additives, and it was found that there is a linear relationship between the solute partition coefficients and the presence of different salt additives. The study suggests that the solute structure and the type of ions influence the solute response to the ionic environment. Additionally, it was observed that the solutes' polar surface area and the solvent-accessible surface area are the essential structural features governing partitioning in aqueous two-phase systems. [ABSTRACT FROM AUTHOR]

Aqueous two-phase systems (ATPSs) are formed when two nonionic polymers, or a single polymer and salt, are mixed in water above a specific concentration, resulting in the emergence of phase separation and the formation of two immiscible aqueous phases. The solvent properties of the aqueous media within the phases of ATPSs rely on the specific composition of the co-solutes and the arrangement of the hydrogen bond network within each phase. Here, we investigate the anion effect of various sodium salts on the enhancement or destabilization of polyethylene glycol (PEG)–salt ATPS formation. Relatively small changes in ATPS ionic composition were shown to result in significant changes in solute partitioning. Additionally, we previously established that the arrangement of hydrogen bonds within the coexisting phases of ATPSs is different, as evidenced by Attenuated Total Reflection—Fourier Transform Infrared (ATR-FTIR) spectroscopic analysis of OH-stretch bands. The hydrogen bond arrangement was shown to abruptly change at concentrations below the threshold of macroscopic phase separation in the ATPSs. Using dynamic light scattering (DLS), we observed a correlation between these abrupt changes in H-bond arrangement and the detection of agglomerate formation in both polymer–polymer and polymer–salt systems. [ABSTRACT FROM AUTHOR]

This review covers the analytical applications of protein partitioning in aqueous two-phase systems (ATPSs). We review the advancements in the analytical application of protein partitioning in ATPSs that have been achieved over the last two decades. Multiple examples of different applications, such as the quality control of recombinant proteins, analysis of protein misfolding, characterization of structural changes as small as a single-point mutation, conformational changes upon binding of different ligands, detection of protein–protein interactions, and analysis of structurally different isoforms of a protein are presented. The new approach to discovering new drugs for a known target (e.g., a receptor) is described when one or more previous drugs are already available with well-characterized biological efficacy profiles. [ABSTRACT FROM AUTHOR]

This short review describes the expansion of the solvatochromic approach utilizing water-soluble solvatochromic dyes to the analysis of solvent features of aqueous media in solutions of various compounds. These solvent features (polarity/dipolarity, hydrogen bond donor ability (HBD acidity), and hydrogen bond acceptor ability (HBA basicity)) vary depending on the nature and concentration of a solute. Furthermore, the solvent features of water (the solvent dipolarity/polarizability and hydrogen bond donor ability) in solutions of various compounds describe multiple physicochemical properties of these solutions (such as the solubility of various compounds in aqueous solutions, salting-out and salting-in constants for polar organic compounds in the presence of different inorganic salts, as well as water activity, osmotic coefficients, surface tension, viscosity, and the relative permittivity of aqueous solutions of different individual compounds) and are likely related to changes in the arrangement of hydrogen bonds of water in these solutions. [ABSTRACT FROM AUTHOR]

Liquid-liquid phase separation underlies the formation of membrane-less organelles inside living cells. The mechanism of this process can be examined using simple aqueous mixtures of two or more solutes, which are able to phase separate at specific concentration thresholds. This work presents the first experimental evidence that mesoscopic changes precede visually detected macroscopic phase separation in aqueous mixtures of two polymers and a single polymer and salt. Dynamic light scattering (DLS) analysis indicates the formation of mesoscopic polymer agglomerates in these systems. These agglomerates increase in size with increasing polymer concentrations prior to visual phase separation. Such mesoscopic changes are paralleled by changes in water structure as evidenced by Attenuated Total Reflection—Fourier Transform Infrared (ATR-FTIR) spectroscopic analysis of OH-stretch bands. Through OH-stretch band analysis, we obtain quantitative estimates of the relative fractions of four subpopulations of water structures coexisting in aqueous solutions. These estimates indicate that abrupt changes in hydrogen bond arrangement take place at concentrations below the threshold of macroscopic phase separation. We used these experimental observations to develop a model of phase separation in aqueous media. [ABSTRACT FROM AUTHOR]

This work presents the first evidence that dissolved globular proteins change the arrangement of hydrogen bonds in water, with different proteins showing quantitatively different effects. Using ATR-FTIR (attenuated total reflection—Fourier transform infrared) spectroscopic analysis of OH-stretch bands, we obtain quantitative estimates of the relative amounts of the previously reported four subpopulations of water structures coexisting in a variety of aqueous solutions. Where solvatochromic dyes can measure the properties of solutions of non-ionic polymers, the results correlate well with ATR-FTIR measurements. In protein solutions to which solvatochromic dye probes cannot be applied, NMR (nuclear magnetic resonance) spectroscopy was used for the first time to estimate the hydrogen bond donor acidity of water. We found strong correlations between the solvent acidity and arrangement of hydrogen bonds in aqueous solutions for several globular proteins. Even quite similar proteins are found to change water properties in dramatically different ways. [ABSTRACT FROM AUTHOR]

Analysis by attenuated total reflection–Fourier transform infrared spectroscopy shows that each coexisting phase in aqueous two-phase systems has a different arrangement of hydrogen bonds. Specific arrangements vary for systems formed by different solutes. The hydrogen bond arrangement is shown to correlate with differences in hydrophobic and electrostatic properties of the different phases of five specific systems, four formed by two polymers and one by a single polymer and salt. The results presented here suggest that the arrangement of hydrogen bonds may be an important factor in phase separation. [ABSTRACT FROM AUTHOR]

Analysis of the partition coefficients of small organic compounds and proteins in different aqueous two-phase systems under widely varied ionic compositions shows that logarithms of partition coefficients for any three compounds or proteins or two organic compounds and one protein are linearly interrelated, although for protein(s) there are ionic compositions when the linear fit does not hold. It is suggested that the established interrelationships are due to cooperativity of different types of solute–solvent interactions in aqueous media. This assumption is confirmed by analysis of distribution coefficients of various drugs in octanol-buffer systems with varied ionic compositions of the buffer. Analysis of the partition coefficients characterizing distribution of variety of drugs between blood and different tissues of rats in vivo reported in the literature showed that the above assumption is correct and enabled us to identify the tissues with the components of which the drug(s) may engage in presumably direct interactions. It shows that the suggested assumption is valid for even complex biological systems. [ABSTRACT FROM AUTHOR]

Background: Simple field tests such as the Timed Up and Go test (TUG) and 30 s Chair Stand test are commonly used to evaluate physical function in the elderly, providing crude outcome measures. Using an automatic chronometer, it is possible to obtain additional kinematic parameters that may lead to obtaining extra information and drawing further conclusions. However, there is a lack of studies that evaluate the test-retest reliability of these parameters, which may help to judge and interpret changes caused by an intervention or differences between populations. Thus, the aim of this study was to evaluate the test-retest reliability of the Timed Up and Go test (TUG) and 30 s Chair Stand test in healthy older adults. Methods: A total of 99 healthy older adults participated in this cross-sectional study. The TUG and the 30 s Chair Stand test were performed five times and twice, respectively, using an automatic chronometer. The sit-to-stand-to-sit cycle from the 30 s Chair Stand test was divided into two phases. Results: Overall, reliability for the 30 s Chair Stand test was good for almost each variable (intraclass correlation coefficient (ICC) >0.70). Furthermore, the use of an automatic chronometer improved the reliability for the TUG (ICC >0.86 for a manual chronometer and ICC >0.88 for an automatic chronometer). Conclusions: The TUG and the 30 s Chair Stand test are reliable in older adults. The use of an automatic chronometer in the TUG is strongly recommended as it increased the reliability of the test. This device enables researchers to obtain relevant and reliable data from the 30 s Chair Stand test, such as the duration of the sit-to-stand-to-sit cycles and phases.