The structural transitions of erythrocyte membranes induced by cyclic AMP.

The generalized structural transitions of erythrocyte membranes induced by cyclic AMP were registered by ESR, fluorescence, freeze-fracture and circular dichroism methods. Two transitions different in nature were revealed. One, which arises at 10-(11)--10-(10) M cyclic AMP, is cooperative and may be considered as a consequence of interaction of cyclic AMP with a receptor. It was calculated that a structural rearrangement in one erythrocyte ghost is induced by three cyclic AMP molecules. As a result of it the membranes are "loosened". The other transition arises at 10-(10)--10-(8) M cyclic AMP and depends on the activity of the protein kinase system. This transition was shown to be non-cooperative and due to phosphorylation of membranous proteins. During this rearrangement the membranes are "stiffened". Both transitions were demonstrated to relate to the membrane integrity.     

On the "receptor" mechanism of the effect of biologically active substances in ultra low concentrations

On the basis of antioxidant (alpha-tocopherol and phenosan potassium salt) and peptide (thyroliberin) effects on the lipid peroxide oxidation (LPO) and lipid structural parameters of the endoplasmic reticulum membranes in wide concentration range (10(-20)-10(-4) mol/l) in vitro the possibility concerning a proposed role of "super-affine" receptors in the mechanism of biologically active substances (BAS) action in ultra low doses (ULD) is discussed. Because these substances modulate investigated processes in the membranes which have not ordinarily receptors the conclusion about availability of non-receptor component in the mechanism of BAS effect in ULD and a low probability of "super-affine" receptor existence has been done.     

当归水提液和醇提液体外抗脂质过氧化和红细胞溶血作用

研究当归水提液和醇提液对小鼠肝组织自发性过氧化酯质分解产物丙二醛（malondialdehyde,MDA）的生成和对红细胞膜脂质过氧化及红细胞溶血作用的影响。采用TBA比色法测定肝组织匀浆MDA生成,分光光度法测定过氧化氢诱导红细胞膜脂质过氧化和溶血。实验分为空白组、对照组、加药组。加药组分为25、50、100和200mg/mL四个浓度组。当归水提液和醇提液均在25～200mg／mL的浓度范围内,能够明显抑制小鼠肝组织匀浆自发性MDA的生成,具有抑制过氧化氢诱导红细胞膜脂质过氧化和溶血的作用,抑制效果随当归水提液和醇提液浓度的增大而逐渐增强,抑制率与药物浓度成良好的量效关系。当归水提液和醇提液具有抗脂质过氧化和红细胞溶血的作用。     

Alpha-Tocopherol-Induced Regulation of Growth and Metabolism in Plants Under Non-stress and Stress Conditions

Alpha-tocopherol (α-Toc) is a member of the vitamin E family and is lipid soluble. Its biosynthesis is by the reaction of isopentyl diphosphate and homogentisic acid in plastid membranes. The putative biochemical activities of tocopherols are linked with the formation of tocopherol quinone species, which subsequently undergo degradation and recycling within cells/tissues. α-Toc plays a key role in a variety of plant metabolic processes throughout the ontogeny of plants. It can maintain the integrity and fluidity of photosynthesizing membranes. It can also neutralize lipid peroxy radicals, consequently blocking lipid peroxidation by quenching oxidative cations. It preserves membrane integrity by retaining membranous structural components under environmental constraints such as water deficiency, high salt content, toxic metals, high/low temperatures, and radiations. α-Toc also induces cellular signalling pathways within biological membranes. Its biosynthesis varies during growth and developmental stages as well as under different environmental conditions. The current review primarily focuses on how α-Toc can regulate various metabolic processes involved in promoting plant growth and development under stress and non-stress and how it can effectively counteract the stress-induced high accumulation of reactive oxygen species (ROS). Currently, exogenous application of α-Toc has been widely reported as a potential means of promoting resistance in plants to a variety of stressful environments.

     

Quaternary phosphonium reversibile inhibitors of cholinesterases of different animals

Quaternary phosphonium compounds were found to be reversible inhibitors of cholinesterases of various animals and showed species-specificity of action depending on the inhibitor structure. It became possible to reveal difference in inhibitory specificity of various preparations of acetylcholinesterases. A difference has been shown in inhibitory parameters of the series of phosphonium toward cholinesterase of visual ganglia of individuals of the squid Berryteuthis magister from different zones of the habitat areal. For the first time, when comparing phosphonium and ammonium isologues - tetrabutyl- and tributylhe-xyl derivatives, it has been shown that they are agents practically similar by the character of anticholinesterase action.     

The association of calmodulin with subcellular fractions isolated from rat liver

Calmodulin associated with rat liver mitochondria has been found to belong to a contaminant membranous fraction which contains different subcellular membranes. The concentration of calmodulin in this fraction is relatively high, about 1.6 micrograms/mg protein, and can not be decreased with EGTA. The calmodulin-rich membranous fraction seems to contain cytoskeletal proteins which could be responsible for the binding of calmodulin.     

Function of DNA Polymerase III in DNA Replication

RECENTLY an in vitro system for DNA replication has been described. This system could be divided into two fractions (A and B) both of which are necessary for proper DNA replication¹. Fraction A, the “soluble” fraction, contains those proteins which do not tightly bind to membranes or native DNA. Fraction B, the “insoluble” fraction, consists of DNA and membranous structures and proteins which are bound to either of them. It was shown that the soluble fraction contains at least one component which is needed at about in vivo concentration¹. Studies of one such component are described in the following.     

AN INTEGRATED MORPHOLOGICAL AND BIOCHEMICAL STUDY OF A PURIFIED PREPARATION OF THE SUCCINATE AND DPNH OXIDASE SYSTEM

Electron micrographs of a purified succinate and DPNH oxidase system prepared from heart muscle reveal that it has a vesicular appearance and is membranous in nature. In keeping with its vesicular appearance is the fact that light scattering by this preparation shows marked changes as the molarity of the suspending medium is altered. Treatment of this preparation with 0.5 per cent deoxycholate solutions removes a large part of the lipide material, which comprises almost half of the dry weight of the preparation. The residue, which still contains the "core" of the cytochrome electron transmitter system, as shown by spectroscopic and enzymatic experiments, is still structured and is membranous in morphological appearance. It is concluded that the enzyme preparation is largely composed of fragmented mitochondrial membranes, and some of the consequences of the localization of the succinate and DPNH oxidase systems in or on these membranes are discussed.     

The association of glycolytic enzymes with cellular and model membranes

This article deals with the binding of glycolytic enzymes with membranous or protein subcellular structures. The representative papers of the last three decades dealing with this matter are reviewed. The studies evidencing the binding of some glycolytic enzymes to insoluble subcellular proteins and membranous structures are presented. It is currently generally accepted that the glycolytic enzymes work in some organisation. Such organisation undoubtedly plays a marked role, although still poorly known, in the regulation processes of glycolysis. From this review, the conclusion emerges that the regulatory ability of the binding of glycolytic enzymes to cellular membranes should be added to the list of well-known mechanisms of post-translational regulation of the glycolytic enzymes. Some of the results presented are the background for the hypothesis that planar phospholipid domains in/on the membrane surface are capable of functioning as binding sites for these enzymes. Such binding can modify the conformation state of the enzymes, which results in changes in their kinetic properties; thus, it may function as a regulator of catalytic activity     

The structural transitions of erythrocyte membranes induced by cyclic AMP

The generalized structural transitions of erythrocyte membranes induced by cyclic AMP were registered by ESR, fluorescence, freeze-fracture and circular dichroism methods. Two transitions different in nature wre revealed. One, which arises at10^?11-10^?10 M cyclic AMP, is cooperative and may be considered as a consequence of interaciton of cyclic AMP with a receptor. It was calculated that a structural rearrangement in one erythrocyte ghost is induced by three cyclic AMP molecules. As a result of it the membranes are “loosened”.The other transition arises at 10^?10-10^?8 M cyclic AMP and depends on the activity of the protein kinase system. This transition was shown to be non-cooperative and due to phosphorylation of membranous proteins. During this rearrangement the membranes are “stiffened”.Both transitions were demonstrated to related to the membrane integrity.     

The membrane-binding segment of dopamine beta-hydroxylase is not an uncleaved signal sequence

Dopamine beta-hydroxylase exists in bovine adrenal medulla chromaffin granules in both soluble and membrane-bound forms. The mechanism by which membranous dopamine beta-hydroxylase is bound to granule membranes has been elusive. Recently, evidence that covalently attached phosphatidylinositol does not serve as an anchor for membranous dopamine beta-hydroxylase was reported (Stewart, L. C., and Klinman, J. P. (1988) J. Biol. Chem. 263, 12183-12186). It was suggested that an uncleaved signal sequence could serve as a mode of attachment for the membrane-bound hydroxylase. Amino-terminal sequence analysis of purified bovine membranous dopamine beta-hydroxylase demonstrates that this form of the enzyme possesses an amino-terminal sequence similar to the soluble enzyme. Additionally, the 75- and 72-kDa bands of membranous dopamine beta-hydroxylase were electrophoretically eluted from a preparative sodium dodecyl sulfate-polyacrylamide gel and sequenced. Both bands had the amino-terminal sequence characteristic of the soluble bovine enzyme. These sequence results eliminate the possibility that an uncleaved signal sequence serves as the membrane anchor.     

Complex dynamic development of poliovirus membranous replication complexes

Replication of all positive-strand RNA viruses is intimately associated with membranes. Here we utilize electron tomography and other methods to investigate the remodeling of membranes in poliovirus-infected cells. We found that the viral replication structures previously described as "vesicles" are in fact convoluted, branching chambers with complex and dynamic morphology. They are likely to originate from cis-Golgi membranes and are represented during the early stages of infection by single-walled connecting and branching tubular compartments. These early viral organelles gradually transform into double-membrane structures by extension of membranous walls and/or collapsing of the luminal cavity of the single-membrane structures. As the double-membrane regions develop, they enclose cytoplasmic material. At this stage, a continuous membranous structure may have double- and single-walled membrane morphology at adjacent cross-sections. In the late stages of the replication cycle, the structures are represented mostly by double-membrane vesicles. Viral replication proteins, double-stranded RNA species, and actively replicating RNA are associated with both double- and single-membrane structures. However, the exponential phase of viral RNA synthesis occurs when single-membrane formations are predominant in the cell. It has been shown previously that replication complexes of some other positive-strand RNA viruses form on membrane invaginations, which result from negative membrane curvature. Our data show that the remodeling of cellular membranes in poliovirus-infected cells produces structures with positive curvature of membranes. Thus, it is likely that there is a fundamental divergence in the requirements for the supporting cellular membrane-shaping machinery among different groups of positive-strand RNA viruses.     

Effect of rhenium complexes with organic ligands on acid resistance of human erythrocytes

The authors showed stabilizing effect of rhenium binuclear cluster compounds with organic ligands on erythrocytic membranes. Estimation of functional condition of cells was studied by a chemical fragility method. Biphase type of rhenium complexes concentration curves stabilization has been found out; the influence of organic radicals and axial substructures (Cl- and Br-) on degree of stabilization has been showed. Different mechanisms of interaction of the investigated complexes with erythrocytic are suggested.     

Enhanced UV sensitivity of Escherichia coli strain uvrA crp]

UV-sensitivity and UV-induced mutability to tryptophan independence has been studied in isogenic crp, cya, crp+, uvrA crp and uvrA crp+ strains of Escherichia coli. crp and cya strains are found to have the same UV-sensitivity as an isogenic wild type strain. UV-sensitivity of uvrA crp strain seems to be one-two orders increased as compared with the sensitivity exhibited by the uvrA - crp+ strain. The yield of UV-induced revertants is slightly higher in crp, cya and uvrA crp strains than in the wild type cells. The existence of cap-dependent inducible error-free repair pathway is supposed due to the data obtained.     

Study of the structural changes in irradiated erythrocyte membranes using 2,6-toluidinonaphthalene sulfonate

It was shown that 2,6-tolyidinonaphthalene sulfonate (2,6-TNS) is localized mainly at the bilayer-water border of the erythrocytic membranes. Under the effect of gamma-radiation the rearrangements occur in the membrane which bring about changes in the distribution of the probe between the membrane and the medium. The lifetime of the excited state of 2,6-TNS after irradiation varies slightly.     

Chemesthesis and the chemical senses as components of a "chemofensor complex"

An important function of the chemical senses is to warn against dangerous biological and chemical agents in the environment. The discovery in recent years of "taste" receptor cells outside the oral cavity that appear to have protective functions has raised new questions about the nature and scope of the chemical senses in general and of chemesthesis in particular. The present paper briefly reviews these findings within the context of what is currently known about the body's chemically sensitive protective mechanisms, including nonsensory processes that help to expel or neutralize threatening agents once they have been encountered. It is proposed that this array of defense mechanisms constitutes a "chemofensor complex" in which chemesthesis is the most ubiquitous, functionally diverse, and interactive chemosensory component.     

Interactions of a Rhodococcus sp. biosurfactant trehalose lipid with phosphatidylethanolamine membranes

Trehalose lipids are an important group of glycolipid biosurfasctants mainly produced by rhodococci. Beside their known industrial applications, there is an increasing interest in the use of these biosurfactants as therapeutic agents. We have purified a trehalose lipid from Rhodococcus sp. and made a detailed study of the effect of the glycolipid on the thermotropic and structural properties of phosphatidylethanolamine membranes of different chain length and saturation, using differential scanning calorimetry, small and wide angle X-ray diffraction and infrared spectroscopy. It has been found that trehalose lipid affects the gel to liquid crystalline phase transition of phosphatidylethanolamines, broadening and shifting the transition to lower temperatures. Trehalose lipid does not modify the macroscopic bilayer organization of saturated phosphatidylethanolamines and presents good miscibility both in the gel and the liquid crystalline phases. Infrared experiments evidenced an increase of the hydrocarbon chain conformational disorder and an important dehydrating effect of the interfacial region of the saturated phosphatidylethanolamines. Trehalose lipid, when incorporated into dielaidoylphosphatidylethanolamine, greatly promotes the formation of the inverted hexagonal HII phase. These results support the idea that trehalose lipid incorporates into the phosphatidylethanolamine bilayers and produces structural perturbations which might affect the function of the membrane.     

The relation of polypeptide hormone structure and flexibility to receptor binding: The relevance of X-ray studies on insulins,glucagon and human placental lactogen

Summary This study is concerned with nondenaturational structural rearrangements of proteins in solution under the influence of physiologically moderate temperatures and salts.Temperature-induced rearrangements are viewed as the reason for breaks in Arrhenius curves of the enzymatic activity. In the cytosol as well as in biological membranes, proteins remain conformationally labile and participate in cooperative structural transitions of membranes. Such transitions are initiated by physiologically moderate temperatures, hormones, salts and aminoacids and affect the functional activity of cell membranes. It is suggested that structural lability of proteins and membranes is of importance in metabolic regulation.It may be said without any exaggeration that a basic objective of biochemistry and biophysics is to find the mechanisms by which coordination of numerous chemical and physicochemical processes along with adaptation to a changing environment can be regulated by the cell. An analysis of a large body of accumulated material and information on this subject leads us to a simple idea; namely, that metabolism is regulated primarily through weak physicochemical interactions. It is weak bonds arising at the sites of contact between effector and regulated macromolecules which serve as a trigger for the regulatory mechanisms of various types. This principle is fundamental for the long range mechanism and for the short-range cytoplasmic allosteric enzyme regulation. In each regulatory act of this type the regulated macromolecule undergoes conformational transition between the states of different functional activity.It is generally recognized^1,2 that in most cases conformational transition is cooperative by nature. However, biopolymers in a cell are an integral part of compact and orderly membraneous phases with active intermolecular interactions. Therefore it is pertinent to inquire whether the elementary act of regulation is necessarily always restricted to one macromolecule or whether there is a possibility of functionally important cooperative transition involving most if not all components of a polymolecular ensembles. We have in mind here the structural long-range effects when local perturbations in the receptor region of the membrane are able to propagate their effects to comparatively large distances. This would occur in a stepwise cooperative transition between two discrete structural states. After we expressed this idea^3,4 we discovered that there had been earlier opinions along such lines⁵ and in recent years similar views have become widely known^6,7,8.Since 1965 our laboratory has been concerned with experimental development of a hypothesis of the membraneous-cooperative-conformational mechanism in the regulation of life processes. We have taken the following path: nondenaturational conformational transitions of proteins in solution rearrangements at the isolated membrane level rearrangements in the intact membrane system of the cells.an invited article.     

Proteolytic Digestion Patterns of "Soluble"and "Detergent-Soluble"Bovine Caudate Nucleus Acetylcholinesterases

Abstract: The structures of purified "soluble"and "detergent-soluble"bovine caudate nucleus acetylcholinesterases were compared by peptide mapping on polyacrylamide gels. The digestion products generated from the two acetylcholinesterases on proteolysis by a given protease ( Staphylococcus aureus V8 protease, α-chymotrypsin, or papain) are remarkably similar as judged from the electrophoretic band patterns. We conclude that the "soluble"and "detergent-soluble"acetylcholinesterases from bovine caudate nucleus share a common evolutionary origin.