[Physical Basis of Functioning of Antifreeze Protein].

Antifreeze proteins, expressed in cold-blooded organisms, prevent ice formation in their bodies, and thus help them to survive in extremely cold winter temperatures. However, the mechanism of action of these proteins is still not clear. In any case, it is not simply a decrease in the temperature of normal ice formation. In this work, investigating the ice-binding protein (a mutant form of the antifreeze protein cfAFP from the spruce budworm Choristoneura fumiferana, which overwinters in needles), we showed that this antifreeze protein does not at all lower the freezing point of water and, paradoxically, increases the melting point of ice. On the other hand, calculations based on the theory of crystallization show that at temperatures of 0° to -30°C ice can only appear on surfaces that contact water, but not in the body of water. These facts suggest a new perspective on the role of antifreeze proteins: their task is not (as it is commonly believed) to bind with nascent ice crystals already formed in the organism and stop their growth, but to bind to those surfaces, on which ice nuclei can appear, and thus completely inhibit the ice formation in supercooled water or biological fluid.

The validity of an experiment testing the influence of acceleration on time dilation using a rotating Mössbauer absorber and a Synchrotron Mössbauer Source.

Three experiments are reviewed, performed (in 2014-2016) at ID18 of ESRF to measure the influence of acceleration on time dilation by measuring the relative shift between the absorption lines of two states of the same rotating absorber with accelerations anti-parallel and parallel to the incident beam. Statistically significant data for rotation frequencies up to 510 Hz in both directions of rotation were collected. For each run with high rotation, a stable statistically significant `vibration-free' relative shift between the absorption lines of the two states was measured. This may indicate the influence of acceleration on time dilation. However, the measured relative shift was also affected by the use of a slit necessary to focus the beam to the axis of rotation to a focal spot of sub-micrometre size. The introduction of the slit broke the symmetry in the absorption lines due to the nuclear lighthouse effect and affected the measured relative shift, preventing to claim conclusively the influence of acceleration on time dilation. Assuming that this loss of symmetry is of first order, the zero value of the relative shift, corrected for this loss, falls always within the experimental error limits, as predicted by Einstein's clock hypothesis. The requirements and an indispensable plan for a conclusive experiment, once the improved technology becomes available, is presented. This will be useful to future experimentalists wishing to pursue this experiment or a related rotor experiment involving a Mössbauer absorber and a synchrotron Mössbauer source.

[Protein Biosynthesis Proofreading Is Closely Associated with the Existence of Factor-Free Ribosomal Synthesis].

Despite protein biosynthesis being studied for decades, some major questions concerning this process are still to be addressed. We elucidate a close connection between proofreading of the emerging amino acid sequence during its normal, elongation factor-dependent ribosomal biosynthesis and the existence of the factor-free synthesis of a polypeptide chain on a ribosome. In this factor-free process, the biological role of proofreading is played by a process opposite to the factor-free attachment of Aa-tRNA to the ribosome, namely, the removal via the same pathway of that Aa-tRNA, which is not complementary to the mRNA codon exhibited by the ribosome.

50+ Years of Protein Folding.

The ability of proteins to spontaneously form their spatial structures is a long-standing puzzle in molecular biology. Experimentally measured rates of spontaneous folding of single-domain globular proteins range from microseconds to hours: the difference - 10-11 orders of magnitude - is the same as between the lifespan of a mosquito and the age of the Universe. This review (based on the literature and some personal recollections) describes a winding road to understanding spontaneous folding of protein structure. The main attention is given to the free-energy landscape of conformations of a protein chain - especially to the barrier separating its unfolded (U) and the natively folded (N) states - and to physical theories of rates of crossing this barrier in both directions: from U to N, and from N to U. It is shown that theories of both these processes come to essentially the same result and outline the observed range of folding and unfolding rates for single-domain globular proteins. In addition, they predict the maximal size of protein domains that fold under solely thermodynamic (rather than kinetic) control, and explain the observed maximal size of "foldable" protein domains.

The Molten Globule Concept: 45 Years Later.

In this review, we describe traditional systems where the molten globule (MG) state has been detected and give a brief description of the solution of Levinthal's paradox. We discuss new results obtained for MG-mediated folding of "nontraditional" proteins and a possible functional role of the MG. We also report new data on the MG, especially the dry molten globule.

Correlates of burnout among professionals working with people with intellectual and developmental disabilities.

BACKGROUND: Although burnout has been recognised as an important stress-related problem among staff working with people with intellectual and developmental disabilities (IDD), literature on the subject is limited yet emerging. The aim of this study is twofold: (1) to evaluate the level of burnout within different professions working with IDD; (2) to examine the association between socio-demographic, professional and organisational characteristics and burnout. METHOD: One hundred ninety-nine professionals working with people with IDD were enrolled in the study (66% response rate). Participants were recruited from several facilities that provide care for people with IDD of all ages, in the Jerusalem area and in other cities in central Israel. The anonymous questionnaires included valid and reliable measures of burnout, socio-demographic variables, professional variables and organisational variables. RESULTS: Participants' mean age was 38.3 years, and most were women. There were no significant differences in burnout levels among the different professionals. Role ambiguity, perceived overload, care-recipient group and job involvement were significant predictors of burnout. The model explained a high percentage (46.8%) of the observed variance. CONCLUSIONS: Most of these predictors are organisational measures. These findings demonstrate that organisational variables are more significantly associated with burnout of staff working with people with IDD than the socio-demographic factors or professional characteristics. Identifying and better understanding the specific factors associated with burnout among professionals working with IDD could facilitate unique intervention programs to reduce burnout levels in staff.

[Evaluation of the Accuracy of Calculation of the Standard Binding Entropy of Molecules from their Average Mobility in Molecular Crystals].

One of the main problems in attempts to predict the binding constants of molecules (or free energies of their binding) is the correct evaluation of configurational binding entropy. This evaluation is possible by methods of molecular dynamics simulation, but these simulations require a lot of computational time. Earlier, we have developed an alternative approach which allows the fast calculation of the binding entropy from summarizing the available data on sublimation of crystals. Our method is based on evaluating the mean amplitude of the movements that are restricted in the bound molecule, e.g., in a crystal, but are not restricted in the free state, e.g., in vapor. In this work, it is shown that the standard entropy of binding of molecules by crystals under standard conditions (1 atm, 25°C) can be assessed rather accurately from geometric and physical parameters of the molecule and the average amplitude of the molecule motions in crystals estimated in our previous work.

[Intersubunit Mobility of the Ribosome].

Ribosomes are ribonucleoprotein nanoparticles synthesizing all proteins in living cells. The function of the ribosome is to translate the genetic information encoded in a nucleotide sequence of mRNA into the amino acid sequence of a protein. Each translation step (occurring after the codon-dependent binding of the aminoacyl-tRNA with the ribosome and mRNA) includes (i) the transpeptidation reaction and (ii) the translocation that unidirectionally drives the mRNA chain and mRNA-bound tRNA molecules through the ribosomal intersubunit space; the latter process is driven by the free energy of the chemical reaction of transpeptidation. Thus, the translating ribosome can be considered a conveying protein-synthesizing molecular machine. In this review we analyze the role of ribosomal intersubunit mobility in the process of translocation.

[Anomalous Kinetics of Amyloidogenesis Suggest a Competition between Oligomers and Fibrils].

Meisl et al. have recently observed an anomalous dependence of the amyloid formation rate on the protein concentration. A novel mechanism of fibril growth has been proposed by Meisl et al. to explain the abnormality; it consists in the fibril-catalyzed initiation of fibril formation with saturation of catalytic sites at high concentrations of substrates. Our article describes an alternative explanation of the anomalous kinetics, assuming that the formation of metastable oligomers competes with fibril formation by decreasing the concentration of free monomers. Oligomers are indeed observed in the course of amyloid formation, but are usually considered as seeds of amyloid fibrils rather as their competitors. However, the oligomers visually detectable by electron microscopy were shown to be close in size to those that can be derived from the anomalous dependence of the amyloid growth rate on the protein concentration, given that the anomaly results from competition between oligomer formation and amyloidogenesis.

Intermediate States of Apomyoglobin: Are They Parts of the Same Area of Conformations Diagram?

Several research teams have reported detection and characterization of various apomyoglobin intermediate states different in their accumulation mode, thus putting a natural question as to proportions of these intermediates. The current report presents spectral properties of sperm whale apomyoglobin studied over a wide range of conditions with the use of circular dichroism and fluorescence techniques. Based on the experimental data, a diagram of apomyoglobin conformational states has been constructed. It shows that though induced by various denaturants, all the observed intermediates belong to one and the same area in the diagram.

Physicians' communication styles as correlates of elderly cancer patients' satisfaction with their doctors.

Physician-patient communication style is of utmost importance to patients with life-threatening diseases. This study identifies the most desired physician communication style by older cancer patients; and examines which of the studied communication styles significantly explains cancer patients' satisfaction with family physicians. A total of 200 older cancer patients, with average age of 75 years, participated in the study, yielding a response rate of 42%. Prospective respondents were randomly selected from the list of cancer patients in the central geographical district of Israel's second largest Health Maintenance Organization fund. Respondents rated their satisfaction with physicians as relatively high. All three communication styles studied were found to be associated with patient's satisfaction. Associations were found between self-rated health, time since the diagnosis of cancer and satisfaction. Women were less satisfied than men with their physicians. Two variables emerged as significant predictors of satisfaction: the physician's caring communication style and patient's gender. Intervention programmes should focus on elevating physicians' awareness of the importance of their communication with cancer patients in general, and of the caring communication style in particular.

[Calculation of mobility and entropy of the binding of molecules by crystals].

A simple method for evaluating a range of molecular movements in crystals has been developed. This estimate is needed to calculate the entropy of binding, in particular in protein-ligand complexes. The estimate is based on experimental data concerning the enthalpy of sublimation and saturated vapor pressure obtained for 15 organic crystals with melting temperatures of 25-80°Ð¡. For this set, we calculated the values of the average range and the corresponding average amplitude of molecular movements in crystals that constituted 0.75 ± 0.14 Å and 0.18 ± 0.03 Å, respectively. The entropy of sublimation calculated based on the average range of molecular movements in crystals was well consistent with the experimental data.

Prevalence of blood type A and risk of vascular complications following transcatheter aortic valve implantation.

OBJECTIVES: To assess the prevalence of blood type A among patients referred for transcatheter aortic valve implantation (TAVI) and whether it is related to vascular complications. BACKGROUNDS: Vascular complications following TAVI are associated with adverse outcomes. Various blood types, particularly type A, have been shown to be more prevalent in cardiovascular diseases and to be related to prognosis. METHODS: The prevalence of various blood types in a cohort of 491 consecutive patients who underwent TAVI was compared with a control group of 6500 consecutive hospitalised patients. The prevalence and predictors of vascular complications and bleeding events were evaluated in the blood type A group and were compared with non-type A patients. RESULTS: The mean age of TAVI patients was 83 ± 6 years, and 40 % were males. Patients were divided into two groups: blood type A (n = 220) and non-type A (n = 271). Type A was significantly more prevalent in the TAVI group than in the control group (45 vs. 38 %, p = 0.023). Compared with the non-type A group, patients with blood type A had more major and fatal bleeding (14.5 vs. 8.1 %, p = 0.027) and more vascular complications (any vascular complication: 24.5 vs. 15.9 % p = 0.016; major vascular complications: 12.3 vs. 7 % p = 0.047). In a multivariable analysis, blood type A emerged as a significant and independent predictor for vascular complications and bleeding events. CONCLUSIONS: Blood type A is significantly more prevalent in TAVI patients than in the general population and is related to higher rates of vascular and bleeding complications.

Synchrotron radiation Mössbauer spectra of a rotating absorber with implications for testing velocity and acceleration time dilation.

Many Mössbauer spectroscopy (MS) experiments have used a rotating absorber in order to measure the second-order transverse Doppler (TD) shift, and to test the validity of the Einstein time dilation theory. From these experiments, one may also test the clock hypothesis (CH) and the time dilation caused by acceleration. In such experiments the absorption curves must be obtained, since it cannot be assumed that there is no broadening of the curve during the rotation. For technical reasons, it is very complicated to keep the balance of a fast rotating disk if there are moving parts on it. Thus, the Mössbauer source on a transducer should be outside the disk. Friedman and Nowik have already predicted that the X-ray beam finite size dramatically affects the MS absorption line and causes its broadening. We provide here explicit formulas to evaluate this broadening for a synchrotron Mössbauer source (SMS) beam. The broadening is linearly proportional to the rotation frequency and to the SMS beam width at the rotation axis. In addition, it is shown that the TD shift and the MS line broadening are affected by an additional factor assigned as the alignment shift which is proportional to the frequency of rotation and to the distance between the X-ray beam center and the rotation axis. This new shift helps to align the disk's axis of rotation to the X-ray beam's center. To minimize the broadening, one must focus the X-ray on the axis of the rotating disk and/or to add a slit positioned at the center, to block the rays distant from the rotation axis of the disk. Our experiment, using the (57)Fe SMS, currently available at the Nuclear Resonance beamline (ID18) at the ESRF, with a rotating stainless steel foil, confirmed our predictions. With a slit installed at the rotation axis (reducing the effective beam width from 15.6 µm to 5.4 µm), one can measure a statistically meaningful absorption spectrum up to 300 Hz, while, without a slit, such spectra could be obtained up to 100 Hz only. Thus, both the broadening and the alignment shift are very significant and must be taken into consideration in any rotating absorber experiment. Here a method is offered to measure accurately the TD shift and to test the CH.

Spaceflight alters host-gut microbiota interactions.

The ISS rodent habitat has provided crucial insights into the impact of spaceflight on mammals, inducing symptoms characteristic of liver disease, insulin resistance, osteopenia, and myopathy. Although these physiological responses can involve the microbiome on Earth, host-microbiota interactions during spaceflight are still being elucidated. We explore murine gut microbiota and host gene expression in the colon and liver after 29 and 56 days of spaceflight using multiomics. Metagenomics revealed significant changes in 44 microbiome species, including relative reductions in bile acid and butyrate metabolising bacteria like Extibacter muris and Dysosmobacter welbionis. Functional prediction indicate over-representation of fatty acid and bile acid metabolism, extracellular matrix interactions, and antibiotic resistance genes. Host gene expression described corresponding changes to bile acid and energy metabolism, and immune suppression. These changes imply that interactions at the host-gut microbiome interface contribute to spaceflight pathology and that these interactions might critically influence human health and long-duration spaceflight feasibility.

Separation of the osmotically driven fusion event from vesicle-planar membrane attachment in a model system for exocytosis.

We demonstrate that there are two experimentally distinguishable steps in the fusion of phospholipid vesicles with planar bilayer membranes. In the first step, the vesicles form a stable, tightly bound pre-fusion state with the planar membrane; divalent cations (Ca++) are required for the formation of this state if the vesicular and/or planar membrane contain negatively charged lipids. In the second step, the actual fusion of vesicular and planar membranes occurs. The driving force for this step is the osmotic swelling of vesicles attached (in the pre-fusion state) to the planar membrane. We suggest that osmotic swelling of vesicles may also be crucial for biological fusion and exocytosis.

Role of osmotic forces in exocytosis: studies of ADH-induced fusion in toad urinary bladder.

Antidiuretic hormone (ADH) treatment of toad urinary bladder activates an exocytotic-like process by which intramembrane particle aggregates are transferred from membranes of elongated cytoplasmic tubules to the luminal-facing plasma membrane. We find that the number of these ADH-induced fusion events, and the number of aggregates appearing in the luminal membrane, are reduced when the luminal bathing medium is made hyperosmotic. As an apparent consequence of the inhibition of their fusion with the luminal membrane, the elongated cytoplasmic tubules become enormously swollen into large, rounded vesicles. These results are consistent with the view that osmotic forces are essential to the basic mechanism of exocytosis.

Parameters affecting the fusion of unilamellar phospholipid vesicles with planar bilayer membranes.

It was previously shown (Cohen, F. S., J. Zimmerberg, and A. Finkelstein, 1980, J. Gen. Physiol., 75:251-270) that multilamellar phospholipid vesicles can fuse with decane-containing phospholipid bilayer membranes. An essential requirement for fusion was an osmotic gradient across the planar membrane, with the vesicle-containing (cis) side hyperosmotic with respect to the opposite (trans) side. We now report that unilamellar vesicles will fuse with "hydrocarbon-free" membranes subject to these same osmotic conditions. Thus the same conditions that apply to fusion of multilamellar vesicles with planar bilayer membranes also apply to fusion of unilamellar vesicles with these membranes, and hydrocarbon is not required for the fusion process. If the vesicles and/or planar membrane contain negatively charged lipids, divalent cation (approximately 15 mM Ca++) is required in the cis compartment (in addition to the osmotic gradient across the membrane) to obtain substantial fusion rates. On the other hand, vesicles made from uncharged lipids readily fuse with planar phosphatidylethanolamine planar membranes in the near absence of divalent cation with just an osmotic gradient. Vesicles fuse much more readily with phosphatidylethanolamine-containing than with phosphatidylcholine-containing planar membranes. Although hydrocarbon (decane) is not required in the planar membrane for fusion, it does affect the rate of fusion and causes the fusion process to be dependent on stirring in the cis compartment.

Black widow spider venom: effect of purified toxin on lipid bilayer membranes.

A purified toxin (the B5 fraction) from black widow spider venom added to the solution on one side of a lipid bilayer membrane interacts irreversibly with the membrane to produce a continuous, linear rise of membrane conductance with time. Conductances greater than 10(-4) reciprocal ohm per square centimeter can eventually be attained without any loss of membrane stability. Membranes treated with toxin are ideally selective for alkali cations over anions and are substantially permeable to calcium ion. These effects of the toxin result from the formation of permanent channels in the membrane of uniform conductance, 3.6 X 10(-10) reciprocal ohm (in 0.1 molar potassium chloride), that remain open almost all the time. Both the divalent cation permeability and the smaller conductances at low pH of toxin-treated membranes suggest that there is negative charge (possibly from carboxyl groups) associated with the channels. We discuss the possible relation of the action of this toxin on lipid bilayer membranes to its ability to stimulate massive transmitter release at the neuromuscular junction and to produce profound morphological changes on tissue cultured neurons.

Micromolar Ca2+ stimulates fusion of lipid vesicles with planar bilayers containing a calcium-binding protein.

Fusion of phospholipid vesicles with a planar phospholipid bilayer membrane that contains a calcium-binding protein appears to mimic the essential aspects of cytoplasmic-vesicle fusion with plasma membranes (exocytosis) in that (i) there is a low basal rate of fusion in the absence of Ca2+, (ii) this basal rate is enormously increased by micromolar (approximately 10 microM) amounts of Ca2+, and (iii) this rate is not increased by millimolar Mg2+. Essential to this process is an osmotic gradient across the planar membrane, with the side containing the vesicles hyperosmotic to the opposite side. Similar osmotic gradients or their equivalent may be crucial for biological fusion events.