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L M Vinokurov

Publications and source records attributed to L M Vinokurov.

16 recordsLinked to original sources
PubMed1998

Green fluorescent protein purification by organic extraction.

Green fluorescent protein (GFP) is widely used as an excellent reporter molecule in biochemistry and cell biology. Some biochemical and immunological assays require high-purity GFP. However, the majority of current procedures for GFP purification include multiple time-consuming chromatography steps with a low yield of the desired product or require tag-containing proteins. An alternative method is described for the GFP purification without affinity extensions using organic extraction yielding a highly homogeneous protein indistinguishable in spectroscopic properties from that purified by previous methods.

1-Butanol↗
PubMed1996

Effect of the ATP level on the overall protein biosynthesis rate in a wheat germ cell-free system.

A sensitive assay which examines the effects of ATP level on the overall activity of a cell-free translation system in a protein synthesis is described. The translational activity of cell-free system was measured in terms of a rate of protein synthesis directed by the 'test' template. The test template encodes a photoluminescent protein, obelin accumulation was determined from the kinetic curves of obelin. The rate of obelin mRNA translation. Time-dependent nucleotide level measurements were conducted throughout the translation processes. It has been shown that the rate of translation decreases exponentially with the decrease of the ATP level. This fall in the overall translation rate is due in part to the mRNA becoming inactive in the translation process. This is not caused by degradation, this mRNA can be restored for translation in a fresh cell-free system by phenol treatment. The reported results provide evidence that the level of ATP unambiguously determines the translational activity of the system.

Adenosine Diphosphate↗
PubMed1983

[Primary structure of the elongation factor G from Escherichia coli. V. Amino acid sequence of the C-terminal domain].

The amino acid sequence of the C-terminal domain of the elongation factor G (EF-G) has been studied. The polypeptide chain of the domain consists of 228 amino acid residues, and contains no tryptophan or cysteine residues. To determine its structure, the peptides obtained as a result of the fragment digestion by staphylococcal glutamic protease, cyanogen bromide cleavage, and tryptic hydrolysis of the fragment modified by maleic anhydride have been analyzed, as well as peptides obtained after hydrolyses of cyanogen bromide fragments with chymotrypsin, thermolysin and trypsin.

Amino Acid Sequence↗
PubMed1983

[Primary structure of the elongation factor G from Escherichia coli. VI. Structure of peptides of cyanogen bromide cleavage of the G-factor molecule].

Peptides obtained as a result of cyanogen bromide cleavage of the G-factor have been studied. All 12 peptides embracing the whole structure of fragment T4 have been isolated. For their amino acid sequence determination, cyanogen bromide peptides have been further cleaved with trypsin, chymotrypsin, thermolysin, staphylococcal glutamic protease and BNPS-skatole. The complete primary structure of 9 from 12 cyanogen bromide peptides has been determined.

Amino Acid Sequence↗
PubMed1983

[Primary structure of the elongation factor G from Escherichia coli. VII. Study of peptides generated during hydrolysis of the T4 fragment by glutamic proteinase from Staphylococcus aureus].

For fragment T4, obtained on limited trypsinolysis of the G-factor, the amino acid sequence embracing 76% of its structure has been determined by analysis of peptides resulting from the fragment T4 cleavage with staphylococcal glutamic protease. These data permitted to assemble into one polypeptide chain 7 out of 12 earlier characterized cyanogen bromide peptides contained in the fragment T4.

Amino Acid Sequence↗
PubMed1983

[Primary structure of the elongation factor G from Escherichia coli. VIII. Structure of tryptic peptides comprising the T4 fragment of limited trypsinolysis of the G-factor].

Products of tryptic hydrolysis of the maleic anhydride modified fragment Th3 from limited thermolytic hydrolysis of the G-factor have been studied. Some short peptides which result from the trypsin action on the native G-factor molecule and belong to the fragment T4 obtained on limited trypsinolysis of the G-factor have been separated and their structure has been studied. As a result amino acid sequence has been determined by tryptic peptides containing 322 amino acid residues of the fragment T4 which makes up about 94% of its polypeptide chain.

Amino Acid Sequence↗
PubMed1983

[Primary structure of the elongation factor G from Escherichia coli. IX. Structure of peptides generated by cyanogen bromide cleavage of the G-factor isolated on thiol-activated sepharose and of the products of the G-factor cleavage at Asp-Pro bonds. Complete primary structure].

The amino acid sequence of cysteine- and cystine-containing peptides resulting from cleavage of the G-factor by cyanogen bromide has been determined. For structure analysis cyanogen bromide peptides were further degradated using trypsin, chymotrypsin, thermolysin, staphylococcal glutamic protease, or limited acid hydrolysis. The products of the G-factor cleavage at Asp-Pro bonds were also studied. The obtained data together with those published earlier permitted to establish the complete primary structure of the elongation factor G. The polypeptide chain consists of 701 amino acid residues and has molecular mass of 77321,46.

Amino Acid Sequence↗
PubMed1983

[Primary structure of calmodulin from the human brain].

The primary structure of calmodulin from human brain has been determined. As compared to calmodulin from bovine brain, it shows three substitutions of the amide----acid type at positions 24, 60, 129, and two acid----amide changes at positions 104 and 135.

Amino Acid Sequence↗
PubMedDate not supplied

[The dependence of stability of the green fluorescent protein-obelin hybrids on the nature of their constituent modules and the structure of the amino acid linker].

Recombinant plasmids containing genes for the green fluorescent protein (GFP) from Aequorea victoria and the photoprotein obelin from Obelia longissima linked in-frame by inserts differing in nucleotide sequences were constructed. The expression of the chimeric genes in Escherichia coli cells resulted in synthesis of the GFP-obelin hybrid proteins. These proteins were purified to homogeneity and subjected to limited trypsinolysis. It was shown that the resistance of GFP-obelin hybrid proteins to trypsin depends on the nature of their constituent modules and the amino acid sequences of linkers between the modules. The kinetics of accumulation of full-length hybrid proteins during the growth of bacterial cells does not depend on the structure of the peptide linkers. Most of the full-length product accumulates in cells in the form of inclusion bodies resistant to endogenous proteases. The soluble fraction of the protein undergoes considerable proteolysis regardless of the linker structure.

Amino Acids↗
PubMedDate not supplied

[Catechol siderophore, produced by thermoresistent strain of Bacillus licheniformis VK21].

Thermophilic and thermoresistant strains of bacilli were screened on a medium containing Chrome Azurol S for producers of siderophores. It was found that the Bacillus licheniformis VK21 strain dramatically increases secretion of the metabolite, a chelator of Fe3+, in response to addition of manganese(II) salts. The growth of the producer on a minimum medium containing MnSO4 under the conditions of iron deficiency is accompanied by the accumulation of a catechol product, the content of which reaches a maximum at the beginning of the stationary growth phase of culture. In the presence of FeCl3, the amount of the catechol product in the medium considerably decreases. The siderophore, called SVK21, was isolated from the cultural medium and purified by reversed phase HPLC, and its siderophore function was confirmed by the test for the restoration of growth of producer cells in a medium containing EDTA. The UV spectrum of the siderophore has absorption maxima at 248 and 315 nm. According to amino acid analysis and NMR spectrometry, the metabolite SVK21 is 2,3-dihydroxybenzoyl-glycyl-threonine. The English version of the paper: Russian Journal of Bioorganic Chemistry, 2003, vol. 29, no. 6; see also http://www.maik.ru.

Bacillus↗