[Increase in glucoamylase productivity of Aspergillus awamori strain by combination of radiating mutagenesis and plasmid transformation methods].

Increase in the level of amylolytic genes activator protein encoded by amyR gene was shown to result in enhancement of glucoamylase productivity of A. awamori strain by 30%. However, the same effect equal to 30% increase can be achieved by introduction of extra copies of gla gene encoding glucoamylase. These two effects were not additive, which gave the possibility to suggest an additional limitation in the egulation mechanism of glucoamylase gene expression in Aspergillus family strains while introducing an additional copies of amyR and gla genes.

[Synthesis of xylanolytic enzymes and other carbohydrases by the fungus Penicillium canescens: transformants with an altered copy number of the gene xlnR and an encoding transcriptional activator].

A fragment of Penicillium canescens genomic DNA carrying the xlnR gene coding for a translational activator of xylanolytic genes was isolated. It was demonstrated that a loss of this function in genetically modified transformants resulted in a drastic decrease in the production of P. canescens major xylanases and had a negative effect on the syntheses of several other extracellular xylanases. An increase in the dose of the xlnR gene elevated the xylanolytic activity as well as the activities of a number of other auxiliary enzymes involved in xylan degradation. The activities of two P. canescens major secreted enzymes--beta-galactosidase and alpha-L-arabinofuranosidase-appeared weakly dependent on the translational activator xlnR.

[Induction of endo-1,4-beta-xylanase and beta-galactosidase in the original and recombinant strains of the fungus Penicillium canescens]. / Induktsiia sinteza éndo-1,4-beta-ksilanazy i beta-galaktozidazy v iskhodnykh i rekombinantnykh shtammakh griba Penicillium canescens.

The induction of the synthesis of secreted enzymes endo-1,4-beta-xylanase (EC 3.2.1.8) and beta-galactosidase (EC 3.2.1.23) in the original and recombinant Penicillium canescens strains has been studied. In all producer strains, the synthesis of these enzymes was induced by arabinose and its metabolite arabitol. The two enzymes differed in the concentration of arabinose required for the induction: the synthesis of beta-galactosidase was most pronounced at 1 mM, whereas maximum synthesis of endo-1,4-beta-xylanase was observed at 5 to 10 mM. An increase in the number of endo-1,4-beta-xylanase copies in the high-copy-number strain of the fungus suppressed the synthesis of beta-galactosidase; the synthesis of endo-1,4-beta-xylanase in the high-copy-number recombinant producing beta-galactosidase was affected to a lesser extent. The amount of the enzymes synthesized did not depend on the saccharide used as a sole source of carbon for growing the mycelium prior to its transfer to the inducer-containing medium.

[Mechanism of overproduction of secreted enzymes in the mycelial fungus Penicillium canescens]. / Mekhanizm superproduktsii sekretiruemykh fermentov u mitselial'nogo griba Penicillium canescens.

The fungus Penicillium canescens strain F178 (VKPM) and its niaD- mutant exhibited an increased capability of synthesizing extracellular enzymes beta-galactosidase (70-80 U/ml) and xylanase (100 U/ml). The synthesis was induced by arabinose and its catabolite, arabitol. A deficiency in arabitol dehydrogenase, leading to arabitol accumulation in the cell, was detected in the chain of reactions of arabinose catabolism. The increased synthesis of beta-galactosidase and xylanase in P. canescens is accounted for by (1) cellular accumulation of the inducer (arabitol) at low concentrations of arabinose in the medium and (2) prevalence of induction over repression.

[Cloning of an endo-1,4-beta-xylanase gene from Penicillium canescens and construction of multicopy strains]. / Klonirovanie gena éndo-1,4-ksilanazy Penicillium canescens i polucheni mul'tikopiinykh shtammov.

The complete gene xylA that encodes endo-1,4-beta-xylanase secreted by Penicillium canescens was cloned and sequenced. The coding region of the gene is separated by eight introns. The protein comprises 302 amino acids of the mature protein and 25 amino acids of the signal peptide. The xylanase of P. canescens belongs to the glycosyl hydrolase family 10. Nucleotide sequences for binding catabolite repression protein CREA and transactivator protein were detected in the promoter region. A set of multicopy strains displaying a seven-eightfold increase in xylanase yield was obtained. The fraction of xylanase in most productive strains amounted to 30-50% of the total secreted protein.

[Molecular cloning of the gene for secreted beta-galactosidase of the filamentous fungus Penicillium canescens]. / Molekuliarnoe klonirovanie gena sekretiruemoi beta-galaktozidazy mitselial'nogo griba Penicillium canescens.

A secreted beta-galactosidase gene from P. canescens was cloned using synthetic oligonucleotide probes and its nucleotide sequence was partially determined. The gene was shown to be present in the genome as a single copy. The deduced primary structure of the signal peptide and the first 25 amino acid residues of the mature protein was established. The structure analysis of the beta-galactosidase gene revealed at least one short intron 64 bp long. The transcription start point was found to be located 89 bp upstream the initiation codon. Some peculiarities of the signal peptide cleavage mechanism are discussed.

[Molecular cloning of cDNA encoding human prointerleukin-6]. / Molekuliarnoe klonirovanie kDNK, kodiruiushchei prointerleikin-6 cheloveka.

The data are presented on the cloning and sequencing of cDNA coding for human interleukin-6. The variability of cDNA proIL-6 cloned from different cellular sources was studied. The variability of cDNA proIL-6 may be expressed as heterogeneity of 5'- and 3'-end sequences of cDNA as well as single base-pair changes.

[Cloning preprolactin cDNA from Pacific salmon in Escherichia coli]. / Klonirovanie kDNK preprolaktina tikhookeanskogo lososia v Escherichia coli.

Prolactin coding mRNA was shown to be a prevalent part of chum salmon (Oncorhynchus keta) pituitary poly(A)-RNA during the spawning period. Clone lambda gtPrk12 was selected from the pituitary cDNA library by means of hybridization with the prolactin probe, and a nucleotide sequence of the insertion was determined and compared to the prolactin coding sequences from rainbow trout and Pacific chinook salmon, which had been published earlier. The sequences compared exhibited a significant homology. The deduced amino acid sequence of the chum salmon prolactin differed from a sequence determined directly in a single position. The prolactin-coding sequence can be used for constructing the bacterial strain producing prolactin.

[Nucleotide sequences encoding glycinin B4 polypeptide of the cultivated soybean Glycine max and its presumed wild-type ancestor Glycine soja in connection with the origin of cultivated soybean]. / Nukleotidnye posledovatel'nosti, kodiruiushchie polipeptid B4 glitsinina kul'turnoi soi Glycine max i ee predpolagaemogo dikogo predka Glycine soja v sviazi s vyiasneniem proiskhozhdeniia kul'turnoi soi.

Nucleotide sequences of cDNAs encoding soybean glycinin B4 polypeptide were compared in three soybean cultivars and two plant introductions of wild soybean Glycine soja. Only two nucleotide substitutions were found in three cultivars G. max, as compared with G. max and G. soja having nucleotide sequences which contain four nucleotide substitutions. These data serve as additional evidence, at molecular level, indicating the origin of G. max from G. soja. On the other hand, the time period required for four nucleotide substitutions' accumulation, as calculated from parameters of molecular evolution of 11S globulins, is much longer than the term having passed after soybean domestication.

[Molecular characteristics of beta-galactosidase secreted by Penicillium canescens]. / Molekuliarnye kharakteristiki sekretiruemoi beta-galaktozidzay Penicillium canescans.

Extracellular beta-galactosidase from P. canescens culture medium was purified by ion-exchange chromatography on DEAE and CM-Sepharose CL-6B and gel filtration. The enzyme active form was shown to be a monomer with a molecular weight of about 120 kDa; the isoelectric point is 6.7 and the sedimentation coefficient is 6.5. In terms of physico-chemical and catalytic properties, the purified enzyme is similar to beta-galactosidases of other fungi of genus Penicillium. The amino acid composition and the NH2-terminal sequence of 24 residues non-homologous to the corresponding sequences of bacterial and yeast beta-galactosidases were determined.

[The parameters of the molecular evolution of the 11S globulins of plants]. / Nekotorye parametry molekuliarnoi évoliutsii 11S-globulinov rastenii.

Nucleotide sequences of cloned cDNA coding for soybean storage protein glycinin and deduced amino acid sequences of basic polypeptides (subfamily II) of glycinin are compared with the amino acid sequences of 11S globulins from other plants. An average number of amino acid substitutions in various evolutionary branches is calculated. A proportional dependence is established between the average number of substitutions per site (the evolutionary distance) and the hypothetical term of divergence of corresponding taxa. The evolution rate of 11S globulins and a term of divergence of the two subfamilies of 11S globulin polypeptides in Fabaceae is estimated.

[Nucleotide sequence of cDNA of glicinin A3 subunits: variability of different species detected by comparative analysis]. / Nukleotidnye posledovatel'nosti kDNK sub''edinits glitsinina podsemeistva A3: razlichnaia vnutrividovaia variabel'nost', vyiavliaemaia pri sravnitel'nom analize.

Several cDNA cloned which code for subfamily A3 subunits (A3B4 and A5A4B3) of soybean storage protein glycinin were analysed by means of restriction mapping and hybrid - selection. The comparison of A3B4 and A5A4B3 subunits cDNA sequences reveals the 90% homology. The nucleotide sequences obtained in the process of this work were compared with those reported about elsewhere, in order to study the interaspecific variability of homologous but not identical storage protein genes of subfamily A3 glycinin subunit. Nucleotide replacements were found to occur 6 times more frequently in A3B4 subunit gene, as compared to A5A4B3 subunit gene (48 replacements against 8).

[Chloroplast DNA cloning in Escherichia coli. II. The properties of the recombinant plasmids bearing the EcoRI fragments of pea chloroplast DNA and the cloning of the DNA sequences with rRNA genes]. / Klonirovanie DNK khloroplastov v Escherichia coli. Soobshchenie II. Nekotorye svoistva rekombinantnykh plazmid, nesushchikh EcoRI-fragmenty DNK khloroplastov gorokha, i klonirovanie posledovatel'nostei DNK s rRNK-genami.

Previously a method of selection of colicine-defective recombinant plasmids by mitomycin C was described. A series of recombinant plasmids (CPS) with various EcoRI-fragments of pea chloroplast DNA has been obtained. This paper describes some properties of cloned fragments replicated in Escherichia coli. The alkali stability of recombinant plasmid DNAs has been demonstrated, indicating the absence of ribonucleotides in their structure. Heterogeneity of chloroplast DNA in nucleotide composition was demonstrated using ultracentrifugation analysis of CPS-plasmid DNAs in CsCl-actinomycin D density gradient. Pea chloroplast rDNA was cloned in recombinant plasmids.

[High-yield isolation of chloroplast DNA from Hordeum vulcare barley protoplasts]. / Vydelenie khloroplastnoi DNK s bol'shim vykhodom iz protoplastov iachmenia Hordeum vulcare.

A fraction of intact chloroplasts free of other cell components in isolated from barley leaf chloroplasts. Instead of mechanical desintegration of plant tissue, the method described includes the cellulysine treatment, thus increasing the yeild of chloroplast DNA. The mean content of DNA per barley chloroplast is found to be 1.10(-4) g. Base composition of barley chloroplast DNA is 39.8 mol.% of G+C. The treatment of chloroplast DNA with restriction endonuclease EcoRI results in the appearance of 17-19 bands under agarose gel electrophoresis.