Effects of inorganic salts on the structural heterogeneity of serum albumin solutions

The formation of protein clusters or a protein-rich phase in undersaturated solutions of biopolymers is considered theoretically on the basis of phase diagrams of a water-protein-salt system. Concentrated (50–200 mg/ml of protein) water-salt solutions of human serum albumin molecules modified by a maleimide spin-label have been studied experimentally using the ESR technique to characterize the significant general features of the system behaviour suggested by the model phase diagrams. The inorganic ion content (NaCl, KSCN, MgCl₂, and CaCl₂) was varied in the range of 10⁻³–4 M. Salt-induced changes in different experimental ESR spin-label parameters based on relations between spectral line widths and amplitudes were determined and compared with the same parameters in salt-free solution. The data on dipole-dipole interactions of spin labels obtained at 77 K and on spin exchange at normal temperatures are indicative of local protein concentration inhomogeneities. The results have been described in terms of salt-induced dissociation of stabilized supramolecular structures in protein solution–protein clusters, liquid-liquid phase transition between the hydration water of clusters and that of individual proteins, and a rise in surface tension which results in protein stabilization. Received: 22 December 1998 / Revised version: 23 August 1999 / Accepted: 24 September 1999     

EEG markers of the disturbed systemic brain activity in hypoxia

Specific rearrangements of the brain bioelectric potential field and the structures where the components (waves) of the main EEG rhythms interact, as well as the stereotactic location and power of the equivalent electrical dipole sources (EEDSs), were studied at various stages of acute experimental hypoxia (breathing for 15–30 min a hypoxic gas mixture containing 8% oxygen in nitrogen). The disrupted intercentral relationships that ensure the formation of the dynamic “morphological equivalent” to support the integrative brain activity, rearrangements of this activity, and the adaptive functions of the whole brain proved to account for partial or complete disintegration of systemic brain activity during acute hypoxia. EEDS tomography showed that EEDSs responsible for the generation of the basic brain rhythmic pattern are normally located in the thalamic structures. At the initial stages of hypoxia, the distribution of the EEDS foci is changed so that the density of EEDSs is increased on the sections that include the hypothalamic region structures, basal nuclei of the forebrain, and the limbic system; the basal, frontal, and medial regions of the temporal lobes of both hemispheres are also involved. With increasing hypoxia, EEDSs appeared in the basal and medial regions of the frontal lobes. At this time, both the surface and deep regions of the frontal lobes of the brain hemispheres are the major targets of the hypoxic effect. At the stages of severe hypoxia, pronounced functional changes in the CNS are observed, including the phenomenon of movement of multiple EEDS foci primarily through the basal and mediobasal regions of the frontal and temporal lobes and in the limbic system structures. Thus, despite the generalized high-amplitude paroxysmal activity that is observed in EEG, a functional disintegration (disruption) of interactions between individual brain regions appears and leads to disturbed regulation of the brain and systemic brain activity. Spatiotemporal EEG markers have been identified that make it possible to assess the individual sensitivity and resistance to hypoxia, as well as the degree of disintegration of his systemic brain activity at different stages of hypoxia.     

Solubility and phase transitions in the water-protein-salt system

The analytical expressions previously obtained for the lines characterizing various phase transitions (sol-gel, liquid-liquid, liquid-solid) that relate the critical molar composition of the water-protein-salt system to individual features of its components (protein charge z, number v of adsorbed ions, and electrolyte activity function Δ) are represented in conventional coordinates of logarithm of protein solubility logS versus salt concentration m ₃. The tendencies of the changes in the positions of phase transition lines in these coordinates as dependent on the parameters v and z and their ratio are determined. The relationship of the salting-out line and the salting-out coefficient with phase transitions is discussed.     

Structural changes in the erythrocyte membrane of the trout Salmo irideus at seasonal acclimatization

Differences of thermostability were studied in red blood cells of the trout Salmo irideus differing in sex and age, as well as structural-dynamic characteristics of erythrocyte membrane proteins at seasonal acclimatization in the interval of reservoir water temperature of 0–19°C. An increase of resistance of erythrocytes to temperature lysis with elevation of the environmental temperature was revealed to be accompanied by a rise of the proteins segmental mobility and a decrease of intermolecular interactions in spectrin-actin cytoskeleton from the data of the ESR spin labeling method. Regulation of erythrocyte stability during acclimation was concluded to occur both changes of the fatty acids chain package at the variations of lipid composition and by changes of the cytoskeleton structural lability. Thereby this provides an increase of the bilayer firmness, on the one hand, while, on the other hand, a rise of elasticity and expansibility of the membrane on the whole, which increases resistance of cells to colloidal-osmotic hemolysis. Changes of concentration of oxygen dissolved in water, which are caused by temperature fluctuations, do not deem to be of crucial importance for structural stability of erythrocytes, as it can be compensated by another mechanism, specifically by changes of affinity of hemoglobin to oxygen.     

Measurement of specific surface energy of protein hydration shell using ESR of spin label

An approach to determining the microscopic surface tension in water-protein matrix using EPR of spin label was developed. The approach in based on the use of viscosity isotherms of correlation time of spin labels bound to the protein macromolecule. Changes in specific surface tension of spin-labeled molecules of serum albumin, hemoglobin, and antibodies were studied depending on protein concentration, the structure of the reporter group, ligand state of the protein, and the presence of salts, water-soluble polymers, and D2O. Possible reasons of microscopic changes in surface tension are discussed.     

Dynamics of heart rate response in sleeping infants exposed to tone stimuli

Heart rate, EEG, and motor responses were recorded following presentation of a series of 6–10 sound stimuli (2.5-s tones of 1000, 4000, and 250 Hz, 70 dB, interstimulus intervals 18–25 s) in neonates aged 9 to 22 weeks during stage 2–3 sleep. The infants (17 of 19) revealed heart rate (HR) changes in response to tone stimuli that consisted in an expanded form of three phases: (1) short-latency (at 1 s after tone presentation) HR deceleration, (2) HR acceleration with a maximum at 3–5 s, and (3) late HR deceleration at 6–9 s of the poststimulus interval. The occurrence rate of the first two phases of cardiac response is relatively constant during a series of stimuli, whereas the likelihood of late HR deceleration is the highest following the first tone presentation and decreases significantly when the stimulus is repeated. Differences in the dynamics and statistical analysis allow a relative independence of all the three response phases to be suggested. The HR acceleration phase is dramatically enhanced in association with the motor response elicited by the sound stimulus. The late HR deceleration phase occurs not only after the first presentation of stimuli, but also when they are repeated if they evoke EEG reaction (vertex potentials) in response to both the beginning and end of the tone sound. Possible mechanisms of the three phases of poststimulus HR changes are: the vagal cardiac reflex associated with the acoustic (adaptive) reflex, activation of sympathetic efferents in combination with the startle reflex, and secondary vagal deceleration of sinus rhythm likely to be associated with the processes of perception (detection) of a “novel” stimulus and to serve as an indirect sign of an orienting reaction.