Department of Genetics of Microorganisms

The Department was organized in 1955 as the Laboratory of Variability of Microorganisms. In 1963 it received its contemporary name. Since 1955 till 1965 the Department had been headed by P. Vizir, Doctor of Biol. Sciences and since 1965 till present time – by B. Matselyukh, Corresponding member of the National Academy of Sciences of Ukraine. The first 10 years of its activity the Department devoted to the studying induced variability and filtering forms of Enterobacteriaceae. In 1961 the Department started research of some streptomycetes which soon became the main object of this studying.

The Department`s main scientific directions are solving of fundamental and applied problems of streptomycetes' genetics: genetic recombination during transformation, conjugation and fusion of protoplasts; chromosome mapping; genetic control of the primary and secondary metabolites biosynthesis; cytoplasmic inheritance; DNA reparation and restriction; genetic engineering; selection of the highly active strains-producers of biologically active substances. The research results are presented herein below in Articles [4, 5] and Monographs [1, 2]. The following may be considered as the Department's main achievements: by means of selection of haploid recombinants and analysis of heteroclones, a circular genetic map of Streptomyces olivaceus VKX was constructed. The map consists of 267 recombination units, on which 46 genes are localized, including 17 reference markers in order to show distances on the map [1, 4]. This map was shown to be similar to the map of S. coelicolor A3(2), which is a classical object of streptomycetes' genetics; similarity can be observed in localization of phenotype-like loci, distances between them and availability of circular symmetry. These facts argue the stability of the gene sequence in streptomycetes throughout the evolution. The next genetic map was constructed for S. griseus 773, the producer of streptomycin. The map included 14 genetic loci with the gene, which codes the antibiotic production (M. Pidhorska). Another map created for S. antibioticus, the industrial producer of oleandomycin, consists of 13 loci and distances between them shown in recombination units (A. Shevchenko, V. Lukyanchuk). Some stages of methionine and adenine biosynthesis in streptomycetes were elucidated. Streptomycetes, unlike the bacteria, were shown to have methylation of homocysteine with the help of cyanocobalamine-dependent transmethylase as the main was in the methionine synthesis (T. Dekhtiarenko). Streptomycetes secret mononucleotides of purines into the cultural liquid, instead of ribosides as yeasts do (Z. Tkachuk).

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Lethal and mutagenic activity of alkylating compounds, UV-rays upon the streptomycetes' spores and mycelium depending on the dosage, pH, physiological state of the cells was studied (I. Danylenko). Ability of nitrosoguanidine to induce mutation most often within the DNA replication fork in the process of growing of synchronized spores was used in order to construct the replication map of streptomycetes where bidirectional chromosome replication and its 60-minute cycle, were shown (M. Mukvych). Six uvs-genes determining reparation of DNA’s pyrimidine dimers, were mapped (V. Lavrinchuk).

A new stage of the Department’s scientific elaborations was related to the research of S. globisporus 1912 strain which is the carrier of two plasmids: pSG1912-1 (10.3 kb), pSG1912-2 (22.4 kb) and a producer of a new anticancer antibiotic. For the lesser plasmid, restriction map was constructed; on its base, pSG1912-4 tsr vector (8.0 kb), which efficiently transforms the recipient strains’ protoplasts and used for gene cloning, was structured (A. Matselyukh). In cooperation with prof. J. Rohr from the Institute of Organic Chemistry, Goettingen University, molecular structure of the new antibiotic, landomycin E was elucidated. The antibiotic consists of the tetracyclic chromophore nucleus of landomycinon A glycosylated with trisaccharide from two residues of D-olivose and one of L-rhodinose. The antibiotic is referred to polyketide compounds of angucycline family and, according to the data gotten by the US National Cancer Institute at Bethesda, in vitro it inhibits growth of 60 different lines of human cancer cells. In collaboration with the scientists from the Institute of Cell Biology, National Academy of Science of Ukraine and Austrian Institute of Cancer Research, Vienna, the mechanism of anticancer activity of landomycin E was clarified. The antibiotic causes apoptosis of cancer cells manifested by cell pressing, chromatin condensation and apoptic corpuscles formation, DNA fragmentation, poly-ADP-ribosyl polymerase splitting induced by caspases 3 and 7, intensive depolarization of mitochondrial membrane, intracellular ADP-pool decreasing and oxidative stress increase. Another important property of landomycin E is its activity against the cancer cells with multidrug resistance (MDR) and a stop of division of cancer cells in G1 phase of cycle, which provides the possibility to combine it with other preparations at clinical practice. In case of intravenous injection to white mice and rats, LD50 of landomycin E was about 74-76 mg/kg of the animal’s weight. It proves average toxicity of the antibiotic comparing to the other anticancer drugs used nowadays. Eims test showed that landomycin E has weak mutagenic activity.

A scientific basis of biotechnology landomycin E production has been developed by us. The growth dynamics of the producer’s culture in laboratory fermenter is characterized by intensive glucose assimilation, dissolved oxygen absorption and accumulation of the biomass and landomycin E during the 48 hours of cultivation. The antibiotic is synthesized by the culture as a primary metabolite and reaches its maximal concentration during the 48 hours of growth.

In collaboration with prof. H. Laatsch from the Institute of Organic and Biomolecular Chemistry, Goettingen University, the chemical structure of a new derivative compound of diketopiperazines was elucidated as L-N-methylphenylalanyl-dehydro-alpha-aminobutyric acid diketopiperazine and its biological activity was first shown as positive regulator of antibiotic biosynthesis and morphogenesis in streptomycetes (B. Matselyukh) [10].

A number of streptomycetes’ endonucleases of restriction and plasmids were isolated and studied; the bank of pSS27 plasmid (13.7 kb) fragments was formed and part of them – sequenced (V. Lukianchuk, V. Polishchuk).

With the help of induced mutagenesis and genetic engineering a number of strains – stable and highly active producers of this antibiotic were obtained. Bacterial, streptomycetes and fungal strains-producers of polymixin B [11], chlortetracycline [12], β-carotene [13], oleandomycin [14] and kanamycin were selected, protected with authorship rights or patents and inculcated into industrial production of the mentioned above antibiotics and vitamin A. For the Bacillus polymyxa and Blakeslea trispora strains – industrial producers of polymixin B and β-carotene, respectively, licenses were sold to foreign manufacturers.

By means of spontaneous and induced mutagenesis the strains of S. globisporus 1912-4Crt and 1912-4Lcp were obtained synthesizing constitutively without illumination beta-carotene and lycopene. These strains represent the biotechnological interest for production of above mentioned carotenoids and their utilization in poultry farming (S. Holembiovska).

The Department’s scholars B. Matselyukh, H. Stryzhkova and A. Stenko are the laureates of Ukraine's State Prize on Science and Technology for the series of essays “Genetics, selection and industrial launch of industrial microorganisms-producers of antibiotics and β-carotene” (1991); B. Matselyukh is a laureate of Zabolotny Prize for his monograph “Genetic maps of microorganisms” (1990).