[Inducible specific lux-biosensors for the detection of antibiotics: construction and main parameters].

Based on Escherichia coli, highly sensitive specific lux-biosensors for the detection of tetracycline and beta-lactam antibiotics, quinolones, and aminoglycosides have been obtained. To make biosensors, bacteria were used that contained fungal plasmids pTetA'::lux, pAmpC'::lux, pColD'::lux, and plbpA'::lux, in which transcription of the reporter Photorhabdus luminescens luxCDABE genes occurred from the inducible promoters of the tetA, ampC, cda, and ibpA genes, respectively. The main parameters (threshold sensitivity and response time) of lux-biosensors were measured.

[SOS-repair--60 years].

This review integrates 60 years of research on SOS-repair and SOS-mutagenesis in procaryotes and eucaryotes, from Jean Weigle experiment in 1953 year (mutagenesis of lambda bacteriophage in UV-irradiated bacteria) to the latest achievements in studying SOS-mutagenesis on all living organisms--Eukarya, Archaea and Bacteria. A key role in establishing of a biochemical basis for SOS-mutagenesis belonges to the finding in 1998-1999 years that specific error-prone DNA polymerases (PolV and others) catalysed translesion synthesis on damaged DNA. This review focuses on recent studies addressing the new models for SOS-induced mutagenesis in Escherichia coli and Home sapiens cells.

[Trigger factor dependent refolding of bacterial luciferases in Escherichia coli cells: kinetics, efficiency and effect of the bichaperone system, DnaKJE-ClpB].

Here were determined the basic parameters of the Tigger Factor (TF) -dependent refolding of thermal inactivated bacterial luciferases. The TF-dependent refolding is less efficient and requires more time than DnaKJE-dependent refolding. The increase in the intracellular concentration of TF leads to an apparent decrease in the level of the thermal inactivated bacterial luciferase refolding.

[Comparative analysis of antirestriction activity of R64 ArdA and ArdB proteins].

Antirestriction proteins ArdA and ArdB are specific inhibitors of the type I restriction-modification enzymes. The transmissible plasmid R64 ardA and yfeB (ardB) genes were cloned in pUC18 and pZE21 vectors. It was shown that the R64 ArdA and ArdB proteins inhibit only restriction activity of the type I restriction-modification enzyme (EcoKI) in Escherichia coli K12 cells.

["Quorum sensing" regulation of lux gene expression and the structure of lux operon in marine bacteria Alivibrio logei].

A group of luminescent strains of marine bacteria Alivibrio logei has been isolated (basins of the Okhotsk, White and Bering Seas). Strains A. logei were shown to be psycrophiic bacteria with an optimal growth temperature of approximately 15 degrees C.

[Effects of the IbpAB AND ClpA chaperones on DnaKJE-dependent refolding of bacterial luciferases in Escherichia coli cells].

The rate and level of DnaKJE-dependent refolding of the thermoinactivated Aliivibrio fischeri luciferase are considerably lower in Escherichia coli ibpA and ibpB mutants than in wild type cells. The rate and level of refolding are lower in E. coli ibpB::kan than in ibpA::kan cells.

[Antimodification activity of the ArdA and Ocr proteins].

The ArdA and Ocr antirestriction proteins, whose genes are in transmissible plasmids (ardA) and bacteriophage genomes (0.3 (ocr)), specifically inhibit type I restriction-modification enzymes. The Ocr protein (T7 bacteriophage) was shown to inhibit both restriction (endonuclease) and modification (methylase) activities of the EcoKI enzyme in a broad range of intracellular concentrations (starting from 10-20 molecules per cell).

[Proteolytic control of expression of Vibrio fischeri lux-operon genes in Escherichia coli cells].

The key elements of the regulatory system activating expression of the lux-operon genes in the sea bacteria Vibrio fischeri are the LuxR protein (an activator oftranscription) and N-(3-oxohexanoyl) L-homoserine lactone (an autoinducer, AI). It is shown that the ATP-dependent proteases ClpXP and Lon take part in the negative control of expression of the lux-operon genes and that AI protects the LuxR protein from proteolysis.

[The C-terminal domain of the Vibrio fischeri transcription activator LuxR is not essential for degradation by Lon protease].

The Vibrio fischer luxICDABEG genes are activated by autoinducer N-(3-oxohexanoyl)homoserine lactone and the LuxR protein. The LuxR contains 250 aa and consists of two domains. The C-domain, that extends from around residue 162 to the C-terminus, is thought to bind lux regulatory DNA and activate transcription of the luxICDABEG genes.

[Antirestriction proteins ardA and Ocr as effective inhibitors of the type I restriction-modification enzymes].

Genes encoding antirestriction proteins (antirestrictases, inasmuch as the antirestriction proteins inhibit the activity of restriction-modification systems, but have no proper enzyme activity, the name antirestrictase is only tentative) are included in the composition of conjugative plasmids (genes ardABC) and some bacteriophages (genes ocr and darA). Antirestriction proteins inhibit of the type I restriction-modification enzymes and thus protect unmodified DNA of plasmids and bacteriophages from degradation. Antirestriction proteins belong to the "protein mimicry of DNA" family: the spatial structure is like the B-form of DNA, and therefore the antirestriction proteins operated on the principle of concurrent inhibition replacing DNA in the complex with the restriction-modification enzyme.

[Antirestriction and antimodification activities of the T7 Ocr protein: effect of mutations in interface].

Antirestriction protein Ocr (bacteriophage T7) is specific inhibitor of the type I restriction-modification enzymes. The bacteriophage T7 0.3 (ocr) gene is cloned in pUC18 vector.

[Mutation clpA::kan in gene encoding the chaperone of Hsp100-family decreases DnaK-dependent refolding efficiency of proteins in Escherichia coli cells].

The rate and level of DnaK-dependent refolding of the thermoinactivated Vibrio fischeri luciferase were considerably lower in clpA mutant (clpA::kan) then in wild type cells. The decline of level of refolding makes progress with the increase of thermoinactivation time of enzyme. The mutation in clpP gene has no influence on kinetic and level of luciferase refolding.

[Induction of oxidative stress and SOS response in Escherichia coli by plant extracts: the role of hydroperoxides and the synergistic effect of simultaneous treatment with cisplatinum].

Plasmids containing a transcription fusion of Escherichia coli katG, soxS, and resA promoters to the Photorhabdus luminescens lux operon (luxCDABE) were constructed. The bioluminescence method of assessing oxidative stress and SOS response in E. coli cells was applied to test the genotoxicity of cisplatinum and vegetable extracts.

[Influence of mutations at genes of global transcriptional regulators on production of autoinducer AI-2 Quorum Sensing in the system of Escherichia coli].

The control of gene expression in response to an increase in the bacterial population density (Quorum Sensing) involves low-molecular-weight signal molecules (autoinducers, AI). AI-2 and synthase LuxS mediating its synthesis are widely distributed in Gram-negative and Gram-positive bacteria. In this work, the data were obtained on the role of global regulators of gene expression in AI-2 synthesis in Escherichia coli cells.

[Host factors in the regulation of the Vibrio fischeri lux operon in Escherichia coli cells].

It has been shown that the chaperonin GroEL, together with GroES co-chaperonin and Lon ATP-dependent protease are involved in the regulation of expression of the Vibrio fischeri lux operon in Escherichia coli cells. The cells of E. coli groE (pF1)- bearing a plasmid with the complete V.

[Antirestriction and antimodification activities of the ArdA protein encoded by the IncI1 transmissive plasmids R-64 and ColIb-P9].

Proteins of the Ard family are specific inhibitors of type I restriction-modification enzymes. The ArdA of R64 is highly homologous to ColIb-P9 ArdA, differing only by four amino acid residues of the overall 166. However, unlike ColIb-P9 ArdA, which inhibits both the endonuclease and the methylase activities of EcoKI, the R64 ArdA protein inhibits only the endonuclease activity of this enzyme.

[Antirestriction activity of the IncI1 transmissive plasmid R64 ArdA protein].

The transmissive plasmid IncI1 R64 contains the ardA gene encoding the ArdA antirestriction protein. The R64 ardA gene locating in the leading region of plasmid R64 has been cloned and their sequence has been determined. Antirestriction proteins belonging to the Ard family are specific inhibitors of type I restriction-modification enzymes.

[The effect of Clp proteins on DnaK-dependent refolding of bacterial luciferases].

A study was made of the refolding of bacterial luciferases of Vibrio fischeri, V. harveyi, Photobacterium phosphoreum, and Photorhabdus luminescens. By reaction rate, luciferases were divided into two groups.

[The effects of the regulatory proteins RcsA and RcsB on the expression of the Vibrio fischeri lux operon in Escherichia coli].

A study was made of the effect of RcsA and RcsB on the Vibrio fischeri lux expression in Escherichia coli. RcsA suppressed the LuxR activity and thereby inhibited expression of the lux genes coding for luciferase and reductase. In osmotic shock, RcsA-RcsB activated lux expression and, consequently, bioluminescence of E.

[Osmotic shock induces expression of Vibrio fischeri lux genes in Escherichia coli cells].

The effect of osmotic shock on the expression of genes in the lux regulon of marine bacteria Vibrio fischeri was studied in cells of Escherichia coli. Bioluminescence of cells was shown to drastically increase, when cells were exposed to osmotic shock at the early logarithmic growth phase. The expression of lux genes induced by osmotic shock is determined by the two-component regulatory system RcsC-RcsB.

[Role of "anti-restriction" motif in functional activity of anti-restriction protein ArdA pKM101 (incN)].

A number of mutant forms of the antirestriction protein ArdA encoded by the ardA gene located in a transmissive IncN plasmid pKM101 have been constructed. Proteins belonging to the Ard family are specific inhibitors of type I restriction--modification enzymes. Single mutational substitutions of negatively charged amino acid residues located in the "antirestriction motif" with hydrophobic alanine, E134A, E137A, D144A, or a double substitution E134A, E137A do not affect the antirestriction activity (Ard) of ArdA but almost completely abolish the antimodification activity (Amd).

[Sequencing and comparative analysis of the lux operon of Photorhabdus luminescens strain ZM1: ERIC elements as putative recombination spots].

The EcoRI chromosomal fragment (6782 bp) containing the lux operon of Photorhabdus luminescens was cloned in pUC18 and completely sequenced. Enteric repetitive intergenic consensus (ERIC), an imperfect palindrome (125-127 bp) characteristics for Enterobacteriaceae genomes, was found in three sites. Strain Zm1 proved to differ in ERIC number and location from strains Hb, Hm, and Hw.

["Quorum sensing", or how bacteria "talk" to each other].

Recent advances in studying the quorum-sensing systems, which regulate gene expression depending on population density, are reviewed. Low-molecular-weight acyl derivatives of L-homoserine lactone (N-AHL) freely diffuse through cell membranes and determine cell-to-cell communications in bacteria. The quorum-sensing systems have first been found to regulate bioluminescence in marine bacteria Photobacterium (Vibrio) fischeri and Vibrio harveyi.

[Alleviation of type I restriction in Escherichia coli K12 in the presence of the arsR gene from pKW301 of Acidiphilium multivorum AIU 301].

A study was made of the antirestriction activity of Acidiphilium multivorum AIU 301 ArsR, a repressor of the ars operon which confers resistance to arsenite and arsenate and is on pKW301. In Escherichia coli, arsR cloned under the control of Plac in a multi-copy vector alleviated restriction of nonmodified lambda DNA by a factor of 120, six times more efficiently than its analogs of conjugal plasmids R64 (incI1) and R773 (incFI). Amino acid sequence analysis showed that the three ArsR proteins have a homologous region of 38 residues, including the antirestriction motif, in their N domains, whereas the motif is in the C domain in the Ard proteins.