Engineering of a target site-specific recombinase by a combined evolution- and structure-guided approach

Site-specific recombinases (SSRs) can perform DNA rearrangements, including deletions, inversions and translocations when their naive target sequences are placed strategically into the genome of an organism. Hence, in order to employ SSRs in heterologous hosts, their target sites have to be introduced into the genome of an organism before the enzyme can be practically employed. Engineered SSRs hold great promise for biotechnology and advanced biomedical applications, as they promise to extend the usefulness of SSRs to allow efficient and specific recombination of pre-existing, natural genomic sequences. However, the generation of enzymes with desired properties remains challenging. Here, we use substrate-linked directed evolution in combination with molecular modeling to rationally engineer an efficient and specific recombinase (sTre) that readily and specifically recombines a sequence present in the HIV-1 genome. We elucidate the role of key residues implicated in the molecular recognition mechanism and we present a rationale for sTre’s enhanced specificity. Combining evolutionary and rational approaches should help in accelerating the generation of enzymes with desired properties for use in biotechnology and biomedicine.     

Features of the Organization of Populations of a Rare Species Cephalaria uralensis (Murr.) Schrad. ex Roem. et Schult. (Dipsacaceae,Magnoliópsida) in the Trans-Volga and Cis-Urals Regions

Biology Bulletin - Abstract—The results of the study of 23 natural cenopopulations of Cephalaria uralensis (Murr.) Schrad. ex Roem. et Schult., a rare subendemic species of the Eastern...     

Polymorphism of genes for xenobiotic metabolism in petrochemical workers

Genetic polymorphism of xenobiotic metabolizing enzymes responsible for individual susceptibility to different environmental factors was examined in a cohort of petrochemical workers occupationally exposed to adverse action of chemical compounds. Molecular genetic analysis of the 1462V mutation in exon 17 of the CYP1A gene demonstrated close similarity between the genotype and allele frequency distribution patterns in the industrial and control groups. No association between the CYP1A polymorphic alleles and genotypes and the duration of service and concomitant diseases was observed. The odds ratio of the disease development in the workers carrying heterozygous CYP1A1 mutant allele was 2.2. Analysis of the STM1 gene polymorphism demonstrated a decrease in the frequency of the homozygous deletion carriers in the workers compared to the control group. There were no substantial differences between the industrial and control groups with respect to the frequencies of rapid and slow acetylator genotypes revealed at the analysis of the NAT2 gene polymorphism. However, considering the concomitant diseases, in the corresponding industrial subgroup a clear trend towards lower frequency of rapid acetylators was demonstrated. In addition, the odds ratio of the disease development for the workers with slow acetylator phenotype was 1.7.     

Paradoxical rescue from ischemic lung injury by inhaled carbon monoxide driven by derepression of fibrinolysis

Carbon monoxide (CO) can arrest cellular respiration, but paradoxically, it is synthesized endogenously by heme oxygenase type 1 (Ho-1) in response to ischemic stress. Ho-1-deficient (Hmox1-/-) mice exhibited lethal ischemic lung injury, but were rescued from death by inhaled CO. CO drove ischemic protection by activating soluble guanylate cyclase and thereby suppressed hypoxic induction of the gene encoding plasminogen activator inhibitor-1 (PAI-1) in mononuclear phagocytes, which reduced accrual of microvascular fibrin. CO-mediated ischemic protection observed in wild-type mice was lost in mice null for the gene encoding PAI-1 (Serpine1). These data establish a fundamental link between CO and prevention of ischemic injury based on the ability of CO to derepress the fibrinolytic axis. These data also point to a potential therapeutic use for inhaled CO.     

Some observations on the effect of Daflon (micronized purified flavonoid fraction of Rutaceae aurantiae) in bancroftian filarial lymphoedema

BACKGROUND: Morbidity management is a core component of the global programme for the elimination of lymphatic filariasis. In a double-blind clinical trial, the tolerability and efficacy of Daflon (500 mg) + DEC (25 mg) or DEC (25 mg) alone, twice daily for 90 days, was studied in 26 patients with bancroftian filarial lymphoedema. RESULTS: None of the patients in either drug group reported any adverse reaction throughout the treatment period (90 days). Haematological and biochemical parameters were within normal limits and there was no significant difference between the pre-treatment (day 0) and post-treatment (day 90) values. The group receiving Daflon showed significant reduction in oedema volume from day 90 (140.6 PlusMinus; 18.8 ml) to day 360 (71.8 PlusMinus; 20.7 ml) compared to the pre-treatment (day 0, 198.4 PlusMinus; 16.5 ml) value. This accounted for a 63.8% reduction in oedema volume by day 360 (considering the pre-treatment (day 0) as 100%). In the DEC group, the changes in oedema volume (between day 1 and day 360) were not significant when compared to the pre-treatment (day 0) value. The percentage reduction at day 360 was only 9%, which was not significant (P > 0.05). CONCLUSION: This study has shown that Daflon (500 mg, twice a day for 90 days) is both safe and efficacious in reducing oedema volume in bancroftian filarial lymphoedema. Further clinical trials are essential for strengthening the evidence base on the role of this drug in the morbidity management of lymphatic filariasis.     

Direct intracardiac injection of umbilical cord-derived stromal cells and umbilical cord blood-derived endothelial cells for the treatment of ischemic cardiomyopathy

The development of new therapeutic strategies is necessary to reduce the worldwide social and economic impact of cardiovascular disease, which produces high rates of morbidity and mortality. A therapeutic option that has emerged in the last decade is cell therapy. The aim of this study was to compare the effect of transplanting human umbilical cord-derived stromal cells (UCSCs), human umbilical cord blood-derived endothelial cells (UCBECs) or a combination of these two cell types for the treatment of ischemic cardiomyopathy (IC) in a Wistar rat model. IC was induced by left coronary artery ligation, and baseline echocardiography was performed seven days later. Animals with a left ventricular ejection fraction (LVEF) of ≤40% were selected for the study. On the ninth day after IC was induced, the animals were randomized into the following experimental groups: UCSCs, UCBECs, UCSCs plus UCBECs, or vehicle (control). Thirty days after treatment, an echocardiographic analysis was performed, followed by euthanasia. The animals in all of the cell therapy groups, regardless of the cell type transplanted, had less collagen deposition in their heart tissue and demonstrated a significant improvement in myocardial function after IC. Furthermore, there was a trend of increasing numbers of blood vessels in the infarcted area. The median value of LVEF increased by 7.19% to 11.77%, whereas the control group decreased by 0.24%. These results suggest that UCSCs and UCBECs are promising cells for cellular cardiomyoplasty and can be an effective therapy for improving cardiac function following IC.     

Development of views on natural focality of plague

The theory of natural focality of human diseases, basically formulated by Academician E.N. Pavlovsky in 1939, invigorated research on plague, the disease whose epidemics had taken millions of lives in the past. Numerous studies in Russia and abroad have provided a great amount of data on the pathogen, its carriers, and vectors, specific features of infection spread and dynamics of activity of natural plague foci. Over a long period, plague was considered to be an obligate transmissible zoonosis. However, recent laboratory experiments and direct observations in natural foci of this infection indicate that plague is probably a zoophilous sapronosis.     

Detection of RNA and specific antibodies against influenza virus A (H5N1) in human samples

Results of inspection 451 persons in the age of from 19 till 58 years and 2 children in the age of till 2 years on presence of specific antibodies and RNA of the infuenza virus A (H5N1) are submitted. RNA of infuenza virus A (H5N1) in researched samples is not revealed. Specific antibodies to a infuenza virus A (H5N1) in a blood serum of 211 patients and donors it is not revealed.