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There is growing concern about the emergence and spread of multidrug-resistant Neisseria gonorrhoeae. To effectively control antibiotic-resistant bacterial pathogens, it is necessary to develop new antimicrobials and to understand the resistance mechanisms to existing antibiotics. In this study, we discovered the unexpected onset of drug resistance in N. gonorrhoeae caused by amino acid substitutions in the periplasmic chaperone SurA and the β-barrel assembly machinery component BamA. Here, we investigated the i19.05 clinical isolate with mutations in corresponding genes along with reduced susceptibility to penicillin, tetracycline, and azithromycin. The mutant strain NG05 (surAmut bamAmut, and penAmut) was obtained using the pan-susceptible n01.08 clinical isolate as a recipient in the transformation procedure. Comparative proteomic analysis of NG05 and n01.08 strains revealed significantly increased levels of other chaperones, Skp and FkpA, and some transport proteins. Efflux pump inhibition experiments demonstrated that the reduction in sensitivity was achieved due to the activity of efflux pumps. We hypothesize that the described mutations in the surA and bamA genes cause the qualitative and quantitative changes of periplasmic chaperones, which in turn alters the function of synthesized cell envelope proteins.

The programmed death-1 receptor, PD-1, is a negative regulator of T-cell activation. The PD-1.3 polymorphism of the PD-1 gene (PDCD1) has been previously shown to be associated with several autoimmune and inflammatory disorders including systemic lupus erythematosus and multiple sclerosis. We examined for the first time PD-1.3 association with another inflammatory disease with strong immune component, IgE-mediated bronchial asthma, its severity and its biochemical markers (total serum IgE and IL-4).PD-1.3 G/A was genotyped by PCR-RFLP analysis using two different populations: Caucasian (492 Russian individuals) and Asian (276 Buryat individuals).We found a significant association of the PD-1.3 polymorphism with IgE-mediated bronchial asthma and total serum IgE level in the Russian population. Combined genotype AA+AG was correlated with risk of developing allergic bronchial asthma (OR = 1.78, 95% CI 1.13-2.78, p = 0.011) and lower concentrations of total serum IgE (p = 0.001) compared with the wild-type genotype GG. However, PD-1.3 was not polymorphic in the Buryat population.PD-1.3 polymorphism of the PD-1 gene (PDCD1) may contribute to the development of allergic asthma in the Russians but not in the Buryats. Our results could be helpful for a better understanding of the effect of this polymorphism on the development of diseases with strong immune components.

Alzheimer's disease is the most prevalent neurodegenerative pathology. According to the amyloid cascade hypothesis, a key event of the Alzheimer's disease pathogenesis is a transition of the β-amyloid peptide (Аβ) from the monomeric form to the aggregated state. The mechanism of Аβ aggregation is intensively studied in vitro, by means of synthetic peptides and various physico-chemical methods allowing evaluation of size, molecular structure, and morphology of the formed aggregates. The paper reviews both the well-known and recently introduced physico-chemical methods for analysis of Аβ aggregation, including microscopу, optical and fluorescent methods, method of electron paramagnetic resonance, electrochemical and electrophoretic methods, gel-filtration, and mass spectrometric methods. Merits and drawbacks of the methods are discussed. The unique possibility to simultaneously observe Аβ monomers as well oligomers and large aggregates by means of atomic force microscopy or fluorescence correlation spectroscopy is emphasized. The high detection sensitivity of the latter method, monitoring the aggregation process in Аβ solutions at low peptide concentrations is underlined. Among mass spectrometric methods, the ion mobility mass spectrometry is marked out as a method enabling to obtain information about both the spectrum of Аβ oligomers and their structure. It is pointed out that the use of several methods giving the complementary data about Аβ aggregates is the best experimental approach to studying the process of b-amyloid peptide aggregation in vitro.Bolezn' Al'tsgeĭmera iavliaetsia naibolee rasprostranennym neĭrodegenerativnym zabolevaniem. Soglasno gipoteze “amiloidnogo kaskada”, kliuchevym sobytiem patogeneza bolezni Al'tsgeĭmera iavliaetsia perekhod b-amiloidnogo peptida (Ab) iz monomernogo sostoianiia v agregirovannoe. Mekhanizm agregatsii Ab intensivno izuchaetsia in vitro s ispol'zovaniem sinteticheskikh peptidov s pomoshch'iu fiziko-khimicheskikh metodov, pozvoliaiushchikh otsenivat' razmer, molekuliarnuiu strukturu i morfologiiu obrazuiushchikhsia agregatov. V obzore rassmotreny kak khorosho izvestnye, tak i nedavno poiavivshiesia fiziko-khimicheskie metody issledovaniia agregatsii Ab, vkliuchaia metody mikroskopii, opticheskie i fluorestsentnye metody, metod élektronnogo paramagnitnogo rezonansa, élektrokhimicheskiĭ i élektroforeticheskiĭ metody, gel'-fil'tratsiiu i mass-spektrometricheskie metody. Obsuzhdaiutsia dostoinstv a i nedostatki metodov. Otmechena unikal'naia vozmozhnost' odnovremenno nabliudat' kak monomery Ab, tak i ego oligomery i krupnye agregaty s pomoshch'iu metodov atomno-silovoĭ mikroskopii i fluorestsentnoĭ korreliatsionnoĭ spektroskopii, a takzhe vysokaia chuvstvitel'nost' poslednego, pozvoliaiushchaia sledit' za protsessom agregatsii v nizkokontsentrirovannykh rastvorakh Ab. Sredi mass-spektrometricheskikh metodov vydelen metod spektrometrii ionnoĭ podvizhnosti/mass-spektrometrii, kotoryĭ, nariadu s opredeleniem spektra oligomerov Ab, pozvoliaet poluchat' informatsiiu ob ikh strukture. Odnovremennoe ispol'zovanie neskol'kikh metodov, pozvoliaiushchikh poluchit' vzaimodopolniaiushchie dannye ob agregatakh Ab, iavliaetsia naibolee optimal'nym éksperimental'nym podkhodom k issledovaniiu protsessa agregatsii b-amiloidnykh peptidov in vitro.

Neisseria gonorrhoeae (gonococcus) is a strict human pathogen, which causes gonorrhea--an infectious disease, whose origin dates back to more than two thousand years. Due to the unique plasticity of the genetic material, these bacteria have acquired the capacity to adapt to the host immune system, cause repeated infections, as well as withstand antimicrobials. Since the introduction of antibiotics in 1930s, gonococcus has displayed its propensity to develop resistance to all clinically useful antibiotics. It is important to note that the known resistance determinants of N. gonorrhoeae were acquired through horizontal gene transfer, recombination and spontaneous mutagenesis, and may be located both in the chromosome and on the plasmid. After introduction of a new antimicrobial drug, gonococcus becomes resistant within two decades and replaces sensitive bacterial population. Currently Ceftriaxone is the last remaining antibiotic for first-line treatment of gonorrhea. However, the first gonococcus displaying high-level resistance to Ceftriaxone was isolated in Japan a few years ago. Therefore, in the near future, gonorrhea may become untreatable. In the present review, we discuss the chronology of the anti-gonorrhea drugs (antibiotics) replacement, the evolution of resistance mechanisms emergence and future perspectives of N. gonorrhoeae treatment.

Interaction of intranuclear β-amyloid with DNA is considered to be a plausible mechanism of Alzheimer's disease pathogenesis. The interaction of single- and double-stranded DNA with synthetic peptides was analyzed using surface plasmon resonance. The peptides represent the metal-binding domain of β-amyloid (amino acids 1-16) and its variants with chemical modifications and point substitutions of amino acid residues which are associated with enhanced neurotoxicity of β-amyloid in cell tests. It has been shown that the presence of zinc ions is necessary for the interaction of the peptides with DNA in solution. H6R substitution has remarkably reduced the ability of domain 1-16 to bind DNA. This is in accordance with the supposition that the coordination of a zinc ion by amino acid residues His6, Glu11, His13, and His14 of the β-amyloid metal-binding domain results in the occurrence of an anion-binding site responsible for the interaction of the domain with DNA. Zinc-induced dimerization and oligomerization of domain 1-16 associated with phosphorylation of Ser8 and the presence of unblocked amino- and carboxy-terminal groups have resulted in a decrease of peptide concentrations required for detection of the peptide-DNA interaction. The presence of multiple anion-binding sites on the dimers and oligomers is responsible for the enhancement of the peptide-DNA interaction. A substitution of the negatively charged residue Asp7 for the neutral residue Asn in close proximity to the anion-binding site of the domain 1-16 of Aβ facilitates the electrostatic interaction between this site and phosphates of a polynucleotide chain, which enhances zinc-induced binding to DNA.

The interaction of the 16-mer synthetic peptide (Aβ16), which represents the metal-binding domain of the amyloid-β with DNA, was studied employing the surface plasmon resonance technique. It has been shown that Aβ16 binds to the duplex DNA in the presence of zinc ions and thus the metal-binding domain can serve as a zinc-dependent DNA-binding site of the amyloid-β. The interaction of Aβ16 with DNA most probably depends on oligomerization of the peptide and is dominated by interaction with phosphates of the DNA backbone.

In order to investigate whether single nucleotide polymorphisms G(+2722)C and 3020insC in CARD15 gene and Asp299Gly in TLR4 gene contribute to atopic bronchial asthma we performed a comparative analysis of alleles and genotypes frequencies of these polymorphisms in Russian patients from Moscow. DNA samples from 283 patients with atopic bronchial asthma and 227 healthy donors were genotyped. There were associations neither of G(+2722)C and 3020insC in CARD15 gene and Asp299Gly in TLR4 gene with asthma nor of markers of CARD15 gene with asthma severity. Haplotype frequency analysis of CARD15 gene polymorphisms did not reveal significant difference between groups. However, a strong association was found between Asp299Gly and asthma severity. Allele Asp of this marker showed association with mild atopic bronchial asthma and allele Gl--with moderate/severe asthma = 0.47, 95% CI [0.24-0.93] i OR = 2.12, 95% CI [1.08-4.18] respectively).

Using two DNA aptamers selectively recognizing anion-binding exosites 1 and 2 of thrombin as a model, it has been demonstrated that their conjugation by a poly-(dT)-linker (ranging from 5 to 65 nt in length) to produce aptamer heterodimeric constructs results into affinity enhancement. The apparent dissociation constant (Kdapp) measured at the optical biosensor Biacore-3000 for complexes of thrombin with the heterodimeric constructs reached minimum values (Kdapp = 0,2-0,4 nМ) which were approximately 30-fold less than for the complexes with the primary aptamers. A photoaptamer heterodimeric construct was designed connecting photoaptamer and aptamer sequences with the poly-(dT)-linker of 35 nt long. The photoaptamer used could form photo-induced cross-links with the exosite 2 of thrombin and the aptamer used could bind to the exosite 1. The measured value of Kdapp for the photoaptamer construct was approximately 40-fold less than that for the primary photoaptamer (5,3 and 190 nM, respectively). Upon exposure to the UV radiation at 308 nm of the equimolar mixtures of thrombin with the photoaptamer construct, the equal yield of the crosslinked complexes was observed at concentrations which were lower by two orders of magnitude than in the case of the primary photoaptamer. It was found that concurrently with crosslinking to thrombin a photo-induced inactivation of the photoaptamer occurs presumably due to formation of the intermolecular crosslinking.

A comparative analysis of allele and genotype distribution of C(-1055)T and R130Q IL13 gene polymorphisms has been performed in Russian patients from the Moscow region. In the study, 283 DNA specimens of atopic bronchial asthma (BA) patients and 227 DNA specimens of healthy donors were used. No association of these markers with ABA development as well as with total IgE concentration has been found. Haplotype frequency analysis did not reveal significant difference between samples. However, significant association ofC(-1055)Tpolymorphism with the disease severity has been revealed (OR = 2.39, 95% confidence interval 1.44-3.98, p = 0.001). Therefore, C(-1055)T polymorphism was shown to be associated with atopic BA progression.

Single nucleotide polymorphisms (SNPs) are now considered as the most perspective and convenient markers for research of genetic bases of multifactorial diseases. Fast development of technologies for exact screening of great volume of genetic information, construction of genomic maps of SNP-markers promotes development of innovative diagnostic systems on the basis of significant SNP for an estimation of individual genetic risk of development of various diseases. In this review the basic aspects of genetics of a diabetes of type 1 and 2 and an opportunity of use SNP as markers for an estimation of individual genetic predisposition to the given diseases are considered.