Cloning of chloroplast psbA and rbcL-genes from cotton Gossipium hirsutum

The fragments of cotton Gossipium hirsutum c.v. 108-f chloroplast genome were cloned in Escherichia coli cells. The cloned psbA and rbcL genes have been selected using the heterologous probes from spinach. The preliminary attempts to clone the complete psbA gene in pUC19 vector failed, probably, due to the toxicity of its product to Escherichia coli cells, and its 5'- and 3'-ends were cloned separately. Reconstruction experiments revealed that while the complete psbA gene was unable to be stably inherited by Escherichia coli cells, its structural part lacking the promoter region could be readily cloned in the bacterial cells.     

The pericentriolar material in Chinese hamster ovary cells nucleates microtubule formation

The structure and function of the centrosomes from Chinese hamster ovary (CHO) cells were investigated by electron microscopy of negatively stained wholemount preparations of cell lysates. Cells were trypsinized from culture dishes, lysed with Triton X-100, sedimented onto ionized, carbon-coated grids, and negatively stained with phosphotungstate. The centrosomes from both interphase and dividing cells consisted of pairs of centrioles, a fibrous pericentriolar material, and a group of virus-like particles which were characteristic of the CHO cells and which served as markers for the pericentriolar material. Interphase centrosomes anchored up to two dozen microtubules when cells were lysed under conditions which preserved native microtubules. When Colcemid-blocked mitotic cells, initially devoid of microtubules, were allowed to recover for 10 min, microtubules formed at the pericentriolar material, but not at the centrioles. When lysates of Colcemid-blocked cells were incubated in vitro with micotubule protein purified from porcine brain tissue, up to 250 microtubules assembled at the centrosomes, similar to the number of microtubules that would normally form at the centrosome during cell division. A few microtubules could also be assembled in vitro onto the ends of isolated centrioles from which the pericentriolar material had been removed, forming characteristic axoneme- like bundles. In addition, microtubules; were assembled onto fragments of densely staining, fibrous material which was tentatively identified as periocentriolar material by its association of CHO can initiate and anchor microtubules both in vivo and in vitro.     

Isolation of calcium pump system and purification of calcium ion-dependent ATPase from heart muscle.

The procedure for the isolation of the highly active fraction of sarcoplasmic reticulum from pigeon and dog hearts is described. The method is based on the partial loading of heart microsomes with calcium and oxalate ions and the precipitation of loaded vesicles in sucrose and potassium chloride concentration gradients. Preparations obtained possess high activity of Ca2+-dependent ATPase and are also able to accumulate up to 10 mumol Ca2+ per mg protein. Purification of sarcoplasmic reticulum membranes is accompanied by a decrease in concentration of cytochrome a+a3 and an increase in the content of [32P]phosphoenzyme. The basic components in "calcium-oxalate preparation" from hearts are proteins with molecular weights of about 100000 (Ca2+-dependent ATPase) and 55000 Calcium-oxalate preparation from pigeon hearts was used for subsequent purification of Ca2+-dependent ATPase. Specific activity of purified enzyme from pigeon hearts is 12-16 mumol Pi/min per mg protein. Enzyme activity of purified Ca2+-dependent ATPase is inhibited by EGTA and is not sensitive to azide, 2,4-dinitrophenol and ouabain. The data obtained demonstrate the similarity of calcium pump systems and Ca2+-dependent ATPases isolated from heart and skeletal muscles.     

Generation and Characterization of Human Interferon-beta Neutralizing Humanized Antibody

Russian Journal of Bioorganic Chemistry - Humanization of antibodies for the development of novel therapeutic agents with low immunogenicity remains a topical problem in modern science. In the...     

Production of the Extracellular Part of the ErbB2 Receptor for the Study of Immunobiologicals

Russian Journal of Bioorganic Chemistry - The extracellular part of the ErbB2 receptor (ecdErbB2), an oncological marker and a target for therapeutic antibodies, has been expressed in eukaryotic...     

Development of the Bispecific Antibody in Fab-scFv Format Based on an Antibody to Human Interferon Beta-1 and Antibody to HER2 Receptor

Russian Journal of Bioorganic Chemistry - The development of new therapies for malignant tumors is an urgent task. Currently, the humanized antibody trastuzumab is considered the “gold...     

Climate Change Increases Drought Stress of Juniper Trees in the Mountains of Central Asia

Assessments of climate change impacts on forests and their vitality are essential for semi-arid environments such as Central Asia, where the mountain regions belong to the globally important biodiversity hotspots. Alterations in species distribution or drought-induced tree mortality might not only result in a loss of biodiversity but also in a loss of other ecosystem services. Here, we evaluate spatial trends and patterns of the growth-climate relationship in a tree-ring network comprising 33 juniper sites from the northern Pamir-Alay and Tien Shan mountain ranges in eastern Uzbekistan and across Kyrgyzstan for the common period 1935–2011. Junipers growing at lower elevations are sensitive to summer drought, which has increased in intensity during the studied period. At higher elevations, juniper growth, previously favored by warm summer temperatures, has in the recent few decades become negatively affected by increasing summer aridity. Moreover, response shifts are observed during all seasons. Rising temperatures and alterations in precipitation patterns during the past eight decades can account for the observed increase in drought stress of junipers at all altitudes. The implications of our findings are vital for the application of adequate long-term measures of ecosystem conservation, but also for paleo-climatic approaches and coupled climate-vegetation model simulations for Central Asia.     

Effect of early sensory experience on the exploratory activity in adult animals

It was previously found that the exploratory activity of adult Wistar rats with their vibrissae cut in the period from 9 to 20 postnatal days was characterized by lower intragroup variability in comparison with control rats [3]. The present study has shown that the earlier limitation of species-specific afferentation (whisker trimming on postnatal days 2–9) does not induce such changes. We conclude that high plasticity of the brain during the early postnatal period provides better adaptation to the deficit of sensory information.     

Development of a Humanized Antibody 5D3Hu against the PRAME Tumor Antigen

Russian Journal of Bioorganic Chemistry - The PRAME antigen, which is a significant target for monoclonal antibodies, is a tumor-specific marker that is active at all stages of tumor cell...     

Random walk analysis of restricted metabolite diffusion in skeletal myofibril systems

The purpose of this work was the development of a basal mathematical model for the diffusion of low-molecular metabolites in a skeletal muscle cell. A three-dimension diffusion of low-molecular particles was simulated by a Monte-Carlo method (random walks of diffusing molecules). The model takes into account the following structural elements: (i) a regular lattice of actin and myosin filaments inside a myofibril; (ii) the membranes of sarcoplasmic reticulum and mitochondria surrounding the myofibrils; (iii) a set of myofibrils inside a skeletal muscle cell. We simulated diffusion of particles in the bulk of intracellular water phase and their reflections from the rigid surfaces of intracellular structures. The model allowed to calculate the apparent coefficients of particle diffusion in the axial and radial directions, Dparallel(app) and Dperpendicular(app), respectively. In accordance with experimental data from literature, the coefficient Dparallel(app) was independent of time. The coefficient of radial diffusion Dperpendicular(app) decreased with time to steady state values similar to that determined by the NMR diffusion spectroscopy methods. The interactions of diffusing particles with thin and thick filaments of myofibrils could explain the decrease in the Dperpendicular(app) value by a factor of 20%. The collisions of particles with myofilaments began to reveal themselves as a gradual decrease in the Dperpendicular(app) value at early stages of diffusion (t1/2 approximately equal to 0.05 microsec). The contribution of particle reflections from the membranes of sarcoplasmic reticulum and mitochondria to the retardation of the radial diffusion was about of 20-30%, depending on porosity of a membranous shield around the myofibril. For conventional sizes of a membranous shield (diameter 2 microm), the interactions of particles with the shield caused a decrease in the Dperpendicular(app) value with a half-time t1/2 approximately equal to 0.5 msec. This time is essentially lower by a factor about of 100 than that found in published NMR measurements. When we considered diffusion of particles inside a cell compartment confined to impermeable membranous shield, the reflection of particles from this shield led the drastic decrease in the radial diffusion coefficient (Dperpendicular(app) --> porportional to when t --> porportional to). This pattern of the Dperpendicular(app)(t) time-course might be expected in the NMR measurements on skeletal muscle tissue where a sarcolemma represents an impermeable shield for ATP and PCr molecules.