Molecular Genetic and Functional Analysis of the PBS72 Plasmid from Bacillus subtilis Environmental Isolates
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RIMENTAL ARTICLES
Molecular Genetic and Functional Analysis of the pBS72 Plasmid from Bacillus subtilis Environmental Isolates A. S. Gurinovicha, * and M. A. Titoka a
Department of Microbiology, Biological Faculty, Belarussian State University, Minsk, 220030 Belarus *e-mail: [email protected] Received April 1, 2020; revised July 7, 2020; accepted July 13, 2020
Abstract—Analysis of the complete nucleotide sequence of the pBS72 plasmid from an environmental strain Bacillus subtilis 72 (102254 bp) was used to characterize the genetic factors responsible for its vertical and horizontal gene transfer. Replicons similar to pBS72 were found to be present in B. subtilis isolated from various natural environments in Pakistan and the Netherlands (99% identity of the Rep proteins). The relaxase of the pBS72 plasmid, which is responsible for its conjugative transfer, was found to belong to the MobL family. A total of 142 open reading frames were revealed in the genome of the pBS72 plasmid, most of which (93) coded the synthesis of hypothetical polypeptides. Among the known proteins of the pBS72 plasmid genes in members of the genus Bacillus, the polypeptides were revealed, which were responsible for the interaction of bacterial cells with the environment, protection from foreign DNA and damage, as well as those involved in the cellular metabolism and carrying out regulatory functions. A system was developed in order to determine the role of the plasmid in activity of environmental and collection B. subtilis strains. In the presence of 13% NaCl, or at pH values of 4.5 and 11, survival of the environmental strain B. subtilis 72 and of the collection strain B. subtilis 168 was 4.33 and 13 times, 1.75 and 1.95 times, and 1.75 and 9.12 times higher, respectively. UV irradiation resulted in 2.25 times higher survival of B. subtilis 72 and 4.5 times higher survival of B. subtilis 168. The environmental strain B. subtilis 72 had more pronounced adaptive properties, which may indicate existence of additional genetic determinants in their chromosome; interaction of their products with the proteins of plasmid origin was probably responsible for higher resistance to increased osmolarity, high and low pH, and UV irradiation. Keywords: plasmid, genome, Rep protein, relaxase, stress resistance DOI: 10.1134/S0026261720060065
Bacteria Bacillus subtilis ubiquitously present in natural environments are able to utilize a broad range of organic and inorganic substrates and secrete biologically active compounds; they are widely employed in biotechnology as producers of enzymes, antibiotics, and stimulants of plant and animal growth (Harwood et al., 2018). In microorganisms of this taxonomic group, the genetic apparatus can be represented by chromosomal and extrachromosomal genes, and the extrachromosomal genetic elements can replicate according to either the rolling circle (RCR type) or the theta mechanism. It should be noted that all described RCR plasmids of B. subtilis belong to the pC194 family, which frequently occurs in gram-pos
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