[Recombinant human insulin: X. Simplification of folding of the biotechnological precursor of recombinant human insulin].

The folding of biotechnological precursor of human insulin precursor was carried out from solubilized inclusion bodies without a preliminary oxidizing or reducing of Cys residues. The inclusion bodies were dissolved in 8 M urea with addition of 10 mM 2-mercaptoethanol. Hydrophobic cell components were removed from the solution by passing through a neutral weakly hydrophobic sorbent, the solution was five times diluted and refolded upon addition of 0.

[Methods of preparation of recombinant proteins-cytokines. IV. Renaturation of recombinant human interleukin-3].

Renaturation of recombinant human interleukin-3 produced as inclusion bodies in the transformed cells of Escherichia coli was studied and optimized. Importance was shown of removing from the protein solution the hydrophobic cellular components causing irreversible aggregation of the protein under renaturation conditions. An effect of pH on the secondary structure of the denatured protein was revealed by CD spectroscopy.

[Preparation of recombinant cytokines. II. Effective method for isolation, purification and renaturation of human recombinant gamma-interferon].

An efficient method for the isolation, purification, and renaturation of human recombinant gamma-interferon from biomass of transformed E. coli cells was developed. It involves the extraction of the protein from the inclusion bodies, preliminary purification of the protein, and three stages of ion-exchange chromatography with an intermediate renaturation between the second and the third stages.

[Recombinant human insulin. VII. Increased efficacy of chromatographic separation based on a principle of bifunctionality].

Retention mechanisms of insulin and deamido[AsnA21] insulin on the bifunctional sorbent Armsphere-C8(PR) in conditions of reversed-phase chromatography (HPLC and ion-pair HPLC) were studied. In accordance with the chemical differences of these proteins, molecular mechanisms of their interaction with silica gel modified with hydrophobic and ion-exchange groups were revealed. The possibility of simultaneous interaction of sorbed proteins with the stationary phase by both mechanisms under conditions of reversed-phase HPLC was demonstrated.

[An effective method of purifying recombinant human tumor necrosis factor alpha].

An efficient and productive isolation method for human recombinant tumor necrosis factor alpha from Escherichia coli cells was developed. The method includes a membrane filtration step, two steps of ion-exchange chromatography, and gel filtration on a Sephadex G-25 column. The target product was obtained with approximately 50% total yield and greater than 95% purity according to PAGE and HPLC.

[Effect of the topography of the signal peptidase site on the effectiveness of secretion of recombinant human granulocyte-macrophage colony-stimulating factor into Escherichia coli periplasm].

Synthesis of an artificial gene encoding the signal peptide of the Yersinia pestis capsule antigen (Caf1) was accomplished. A set of plasmids coding for hybrid proteins in which a modified sequence of the Caf1 signal peptide is connected to the amino acid sequence of the mature granulocyte-macrophage colony stimulating factor (GM-CSF) were constructed. Topography of the cleavage site of signal proteases was studied.

[Genetic engineering of human insulin. IV. Development and optimization of an analysis system using reversed phase high pressure liquid chromatography].

The effectiveness of the RP HPLC application for the step-by-step analysis of the recombinant insulin production was studied. Properties of a number of commercial and experimental columns in different chromatographic conditions were considered. A three-dimension optimization of selectivity and resolution versus pH and ion strength was carried out.

[Separation of recombinant proteins by chromatography on vesicular sorbents].

Vesicle chromatography, a recently developed method for separation of biomolecules, uses the vesicular packing (VP) material (clusters of microcapsules derived from plant cells), which was tested with respect to its application for the recombinant protein separation. Since VP has a well-defined separation limit, biomolecules are distributed in two separate peaks: large molecules are excluded and small molecules permeate through cell walls into the empty cell lumen. Recombinant proteins frequently form oligomers, which differ from monomers not only in size but also chemically and biologically.

[Genetically engineered human insulin. 1. HPLC in analyzing products of basic production stages].

Application of some variants of HPLC for the step-by-step analysis of recombinant human insulin production was studied. Chromatographic columns with commercial and specially developed supports for size-exclusion, ion-exchange and reverse phase HPLC were used. Effective combinations of the chromatographic techniques for analysis of products and intermediates at every technological step were found and used for production of insulin.

[High performance liquid chromatography of nucleotides. Major methods and their development].

The separation of mono- and oligonucleotides possibilities by means of high performance ion-exchange, reversed-phase, so-called "ion-pair" and adsorption chromatography are studied. The influence of the eluent composition (solvent, salt) and pH on the retention, selectivity and resolution in reversed-phase and ion-exchange chromatography is investigated. The model of the hydrophobic-pair ion-exchange mechanism of ion-pair chromatography is considered.

[Effect of glutamate on the trace element content in the mitochondria of the liver, heart and kidneys of rats with acute and chronic hypoxia].

Acute hypoxia ("height" 8000 metres, 60 minutes) decreases concentration of copper, zinc and magnesium in mitochondria of the liver, kidney, that of copper and magnesium decreases in the heart. Chronic hypoxia ("height" 6000 metres, 6 hours a day for 2 weeks) also causes a decrease in the concentration of copper, zinc and magnesium in mitochondria of the liver, kidney and that of copper and magnesium--in the heart. Glutamic acid (acute and chronic hypoxia) increases the level of copper, zinc and magnesium in mitochondria of the liver, heart and kidney of albino rats which is close to its values in the normal animals.