Vol 42, No 5 (2016)

A-factor, or (2S, 3R)-2-isocapryloyl-3-hydroxymethyl-γ-butirolactone, has been described by A.S. Khokhlov with coworkers and is one of the first studied autoregulators in prokaryotes. A-Factor structure has been confirmed by synthesis. It has been established that actinobacteria produce many substances with structural features closely related to the A-factor, regulating the development of Streptomyces griseus; particularly, they contain γ-bytirolactone, a hydroxymethyl group at position 2, and a fatty acid residue at position 3. These autoregulators are closely related not only in terms of their structure, but also their function, that is, regulation of processes of morphological differentiation, spore formation, and biosynthesis of secondary metabolites, including various antibiotics. This provides grounds for calling them A-factor-like regulators, or gamma-butyrolactones (GBLs), as they are often abbreviated. Structures of 21 natural autoregulators of the group isolated from representatives of eight streptomyces species have been established. Reference strains were used to demonstrate that A-factor regulators are typical of many species of the Actinomycetales order and are specific for these bacteria. They have been described in many species of Streptomyces,Actinomyces, Nocardia, Amycolatopsis, and Micromonospora. Autoregulators exhibit cross-effects with respect to reference strains of various species producing them. Presumably, biosynthesis of A-factor regulators is performed according to a common mechanism starting from fatty acid residues and glycerol as initial metabolites and involving iso-beta-ketoacid; the latter one is cyclized using a molecule of oxidized glycerol forming a nonsaturated gamma-lactone through decondensation at position 2 of oxidized glycerol, which is followed by reduction to A-factor. The first stage of biosynthesis is, supposedly, performed by the product of afsA gene; AfsA is the key enzyme in A-factor biosynthesis. AfsA protein homologues have been found in various streptomyces species. Molecular and genetics studies of A-factor-like autoregulators of S. griseus and some other streptomyces species allowed deciphering a regulatory cascade resulting in morphological differentiation and biosynthesis of secondary metabolites under the effect of nanomolar concentrations of the autoregulators. The action of the A-factor starts with its binding to the A-factor receptor protein (ArpA), which represses the promotor of the target gene. ArpA comprises two domains: N-terminal DNA-binding domain and the A-factor-binding C-terminal domain. ArpA protein binds to the adp4 gene, but DNA is depressed upon A-factor–ArpaA complex formation. This results in transcription of adpA gene encoding a transcription activator AdpA, the central regulator of A-factor regulatory cascade. AdpA amplifies the signal of A-factor, acting as a pleiotropic activator of transcription of at least 72 genes, particularly, the spore formation genes and genes of streptomycin biosynthesis in S. griseus. Altogether, genes activated by AdpA protein form the AdpA regulon. The AdpA-binding consensus DNA sequence has been established. According to their structure, the proteins can be grouped into a larger subfamily of the AraC/XylS family. Study of A-factor-like regulators is of topical interest for both theoretical and practical needs of antibiotic production development.

The minireview is dedicated to recently described enhancer RNAs (eRNAs), which are products of enhancer transcription. Features and putative functions of eRNAs are considered. The potential of eRNAs as therapeutic targets is discussed.

A tritium-labeled synthetic LKEKK pentapeptide corresponding to the sequences 16–20 of human thymosin-α₁ and 131–135 of human interferon-α₂ was obtained with a specific activity of 28 Ci/mmol. [³H]LKEKK was found to bind with high affinity (K_d 3.7 ± 0.3 nM) to the membranes isolated from epithelial cells of rat small intestinal mucosa. The trypsin treatment of the membranes did not affect the binding, thus supporting the nonprotein nature of the peptide receptor. The binding of the labeled peptide was inhibited by unlabeled thymosin-α₁, interferon-α₂, and cholera toxin B subunit (K_i 4.2 ± 0.4, 3.5 ± 0.3, and 4.7 ± 0.3 nM respectively). The pentapeptide did not affect the adenylate cyclase activity within the concentration range of 1–1000 nM.

Human biosynthetic insulin is a polypeptide hormone that plays an important and essential role in control of the level of carbohydrate, protein, and fat metabolism in the blood. Human pancreatic insulin was labeled with ^99mTc to form a new radiopharmaceutical with a labeling yield of 99 ± 1% under optimum conditions: 0.1 mL insulin, pH 7, 25 μg stannous chloride, 1 mL (19 mCi) of pertechnetate, room temperature, and 10 min reaction time. The ^99mTc–insulin complex was examined using paper chromatography, ITLC, electrophoresis, and HPLC. In addition, in vitro and in vivo study of ^99mTc–insulin complex was performed at different time intervals.

Coumarin and heterocyclic compounds incorporating urea have clinical applications as antiepileptics, diuretics, and antiglaucoma agents due to their carbonic anhydrase inhibitory properties. We investigated inhibition of carbonic anhydrase I and II with a series of coumarylthiazole derivatives containing urea/thiourea groups. All the investigated compounds exhibited inhibitory activity on both hCA I and hCA II, with 1-(3-chlorophenyl)-3-(4-(2-oxo-2H-chromen-3-yl)thiazol-2-yl)urea being the strongest inhibitor. Structure–activity relationship study showed that most of urea derivatives were more inhibiting for hCA I and hCA II than thiourea derivatives. The electron-withdrawing groups at the phenyl ring increased the inhibitory activity compared to electron-donating groups.

β-Galactosidases (β-gals) are a wide family of glycosyl hydrolases thought to be involved in the metabolic recycling of galactolipids, glycoproteins, and cell wall polysaccharides in plants. A full-length cDNA, designated STBG2, was isolated and cloned from tomato (Solanum lycopersicum L. cv. Falcato) by reverse transcription–polymerase chain reaction. The cDNA was 2996 bp in length and encoded a typical β-gal protein, designated SlGal2, consisting of 892 residues. A comparison of SlGal2 with its counterpart isoform in the cultivar Rutgers revealed that SlGal2 had identical residues, except for 44 amino acids. Noteworthy, 42 different residues were located in a limited area in the middle of the protein. An alignment of SlGal2 with other plant β-gals clearly showed the existence of a non-conserved short polypeptide, as a connector between the N- and C-terminal domains. A deeper bioinformatic analysis referring to recent experimental findings strongly suggested different roles for the N- and C-terminal domains. As a result, the N-terminal domain is responsible for the catalytic activity and the C-terminal domain is responsible for the stability and the substrate binding of the enzyme. In addition, a substantial difference in physiochemical characteristics of similar β-gals was found to be in their isoelectric points. In conclusion, the differential role of C-terminal domains and also the significant differences in isoelectric points provided insights into the unknown mechanism of substrate specificity of plant β-gals, which in turn will help in protein engineering studies.

Convenient synthetic schemes being suitable for the preparation of aminooxy-Sepharose and hydroxylamine-containing Toyopearl with capacity of 0.35–0.015 mmols of H₂NO-groups per mL gel, including the adsorbent with easily cleavable disulfide bond in the linker are developed.

A novel series of indole based benzofuran derivatives has been synthesized under microwave irradiation and conventional conditions. The structures of the compounds were established on the basis of ¹HNMR, ¹³C NMR, IR and mass spectral data. The analogues were evaluated for their in vitro antimicrobial activity against two gram-positive bacteria, two gram-negative bacteria and two fungal strains. The same series was screened for in vitro antioxidant activity against 2,2-diphenyl-1-picrylhydrazyl (DPPH). Most of the title compounds exhibited promising antimicrobial and antioxidant activities.