Biochemistry (Moscow) (v.77, #4)

In this review, the development of understanding of the biological functions of carnosine is briefly discussed. Carnosine was first described as a component of meat in 1900 by V. S. Gulevitch. Changes in the concepts of the role of carnosine in metabolism are followed starting from the early suggestion that it is the end product of protein degradation to the modern ideas based on demonstrating its specific involvement in intracellular signaling regulation in excitable tissue cells. The discovery of the ability of carnosine to regulate expression of early response genes broadens the concept about carnosine as a cellular peptide regulator. The first attempts for application of carnosine in sport and medical practice are described.
Keywords: carnosine; pH buffer; reactive oxygen species; intracellular signaling; neurodegenerative diseases

Transformation of macrophages into foam cells is traditionally considered in the context of atherogenesis, because lipid accumulation is believed to be a consequence of uptake of oxidized low density lipoproteins (oxLDL) through scavenger receptors (SR) of macrophages. However, an excessive uptake of oxLDL is recently shown to trigger compensatory mechanisms of cholesterol elimination from macrophages. Maintaining the lipid homeostasis in macrophages is mediated by regulation of a system of lipid sensors, which is reprogrammed under conditions of inflammation leading to formation of foam cell phenotype without involvement of SR. The increase in the inflammatory potential on macrophage polarization into the M1 phenotype is associated with suppression of LXR and PPAR, their target genes, induction of expression of genes responsible for fatty acid and cholesterol metabolism controlled by SREBP1c and SREBP2, proteins associated with lipid inclusions, macropinocytosis activation, secretion of LXR and PPAR endogenous ligands, and development of apoptosis. In this review the role of foam cells in development and resolution of acute inflammation, mechanisms of their formation from macrophages infected by some bacterial and virus pathogens causing chronic inflammation, and the significance of LXR and PPAR as therapeutic targets in chronic infectious and inflammatory diseases are also discussed.
Keywords: macrophage/foam cell; inflammation; lipid sensors; endogenous ligands PPAR and LXR; macrophage infections

The level of acute-phase serum amyloid A (SAA) protein in human blood dramatically grows in cancer, often at its early stage, when acute inflammatory signs are not observed. This fact was registered both by immunochemistry and by proteomics methods in different common cancers, such as lung, ovarian, renal, uterine, and nasopharyngeal cancer and in melanoma. It was proposed that SAA is produced by liver in such cases, as in inflammation, high levels of SAA being a part of nonspecific response to tumor. However, that was not always true, because, in many cancers, the protein of interest is produced directly by cancer cells. What is the biological significance of this observation? What preferences do cancer cells obtain due to SAA overexpression? Recent data on melanoma patients have shown that serum amyloid A is able to stimulate immunosuppressive neutrophils to produce interleukin-10 cytokine that suppressed cell immunity. The ability of cancer cells to produce SAA that is acquired during cancer mutagenesis is likely to enhance their resistance to T-cell immunity due to activation of immunosuppressive granulocytes.
Keywords: acute-phase serum amyloid A (SAA); biomarker; proteomics; cancer; malignancy; myeloid derived suppressor cell (MDSC)

Leader sequences of eukaryotic mRNA can be simultaneously bound to initiating 80S ribosome and 40S ribosomal subunit by E. A. Sogorin; N. E. Shirokikh; A. M. Ibragimova; V. D. Vasiliev; S. Ch. Agalarov; A. S. Spirin (342-345).
Binding of mRNA leader sequences to ribosomes was studied in conditions of a cell-free translation system based on wheat germ extract. Leader sequence of TMV mRNA (the so-called omega-RNA sequence) was able to bind simultaneously 80S ribosome and 40S ribosomal subunit. It was found that nucleotide substitutions in omega-RNA resulting in destabilization of RNA structure have no effect on the complex formation with both 80S ribosome and 40S ribosomal subunit. Leader sequence of globin mRNA is also able to form a similar joint complex. It is supposed that the ability of mRNA leader sequences to bind simultaneously 80S ribosome and 40S subunit is independent of leader nature and may reflect previously unknown eukaryotic mechanisms of translation initiation.
Keywords: omega-RNA; initiation of protein biosynthesis; ribosomal complexes; electron microscopy of ribosomes

Influence of centrin 2 on the interaction of nucleotide excision repair factors with damaged DNA by Y. S. Krasikova; N. I. Rechkunova; E. A. Maltseva; C. T. Craescu; I. O. Petruseva; O. I. Lavrik (346-353).
We have examined the influence of centrin 2 (Cen2) on the interaction of nucleotide excision repair factors (XPC-HR23b, RPA, and XPA) with 48-mer DNA duplexes bearing the dUMP derivative 5-{3-[6-(carboxyamidofluores-ceinyl)amidocapromoyl]allyl}-2′-deoxyuridine-5′-monophosphate. The fluorescein residue linked to the nucleotide base imitates a bulky lesion of DNA. Cen2 stimulated the binding and increased the yield of DNA adducts with XPC-HR23b, a protein recognizing bulky damages in DNA. Stimulation of the binding was most pronounced in the presence of Mg2+ and demonstrated a bell-shaped dependence on Cen2 concentration. The addition of Cen2 changed the stoichiometry of RPA-DNA complexes and diminished the yield of RPA-DNA covalent crosslinks. We have shown that Cen2 influences the binding of RPA and XPA with DNA, which results in formation of additional DNA-protein complexes possibly including Cen2. We have also found some evidence of direct contacts between Cen2 and DNA. These results in concert with the literature data suggest that Cen2 can be a regulatory element in the nucleotide excision repair system.
Keywords: nucleotide excision repair; centrin 2; photoaffinity modification; binding of repair protein factors with DNA

Programmed cell death in plants: Protective effect of tetraphenylphosphonium and tetramethylrhodamine cations used as transmembrane quinone carriers by L. A. Vasil’ev; D. B. Kiselevsky; E. V. Dzyubinskaya; A. V. Nesov; V. D. Samuilov (354-361).
Tetraphenylphosphonium (TPP+) and tetramethylrhodamine ethyl ester (TMRE+) cations used as transmembrane carriers of ubiquinone (MitoQ) and plastoquinone (SkQ, SkQR) in mitochondria prevented at nanomolar concentrations the chitosanor H2O2-induced destruction of the nucleus in epidermal cells of epidermis isolated from pea leaves. The protective effect of the cations was potentiated by palmitate. Penetrating anions of tetraphenylboron (TB) and phenyl dicarbaundecaborane also displayed protective effects at micromolar concentrations; the effect of TB was potentiated by NH4Cl. It is proposed that the protective effect of the penetrating cations and anions against chitosan is due to suppression of the generation of reactive oxygen species in mitochondria as a result of the protonophoric effect of the cations plus fatty acids and the anions plus NH 4 + . Phenol was suitable as the electron donor for H2O2 reduction catalyzed by horseradish peroxidase, preventing the destruction of cell nuclei. The penetrating cations and anions, SkQ1, and SkQR1 did not maintain the peroxidase or peroxidase/oxidase reactions measured by their suitability as electron donors for H2O2 reduction or by the oxidation of exogenous NADH.
Keywords: penetrating ions; tetraphenylphosphonium; tetramethylrhodamine ethyl ester; tetraphenylboron anion; phenyl dicarbaundecaborane anion; cell nucleus destruction; protective effect; plant epidermal cells; chitosan; CN

Influence of oxidative and nitrosative stress on accumulation of diphosphate intermediates of the non-mevalonate pathway of isoprenoid biosynthesis in corynebacteria and mycobacteria by V. Yu. Artsatbanov; G. N. Vostroknutova; M. O. Shleeva; A. V. Goncharenko; A. I. Zinin; D. N. Ostrovsky; A. S. Kapreliants (362-371).
Artificial generation of oxygen superoxide radicals in actively growing cultures of Mycobacterium tuberculosis, Myc. smegmatis, and Corynebacterium ammoniagenes is followed by accumulation in the bacterial cells of substantial amounts of 2-C-methyl-D-erythritol-2,4-cyclodiphosphate (MEcDP) — an intermediate of the non-mevalonate pathway of isoprenoid biosynthesis (MEP) — most possibly due to the interaction of the oxygen radicals with the 4Fe-4S group in the active center and inhibition of the enzyme (E)-4-oxy-3-methylbut-2-enyl diphosphate synthase (IspG). Cadmium ions known to inhibit IspG enzyme in chloroplasts (Rivasseau, C., Seemann, M., Boisson, A. M., Streb, P., Gout, E., Douce, R., Rohmer, M., and Bligny, R. (2009) Plant Cell Environ., 32, 82–92), when added to culture of Myc. smegmatis, substantially increase accumulation of MEcDP induced by oxidative stress with no accumulation of other organic phosphate intermediates in the cell. Corynebacterium ammoniagenes, well-known for its ability to synthesize large amounts of MEcDP, was also shown to accumulate this unique cyclodiphosphate in actively growing culture when NO at low concentration is artificially generated in the medium. A possible role of the MEP-pathway of isoprenoid biosynthesis and a role of its central intermediate MEcDP in bacterial response to nitrosative and oxidative stress is discussed.
Keywords: non-mevalonate pathway; nitrosative stress; mycobacteria

Properties of recombinant ATP-dependent fructokinase from the halotolerant methanotroph Methylomicrobium alcaliphilum 20Z by S. Y. But; O. N. Rozova; V. N. Khmelenina; A. S. Reshetnikov; Y. A. Trotsenko (372-377).
In the cluster of genes for sucrose biosynthesis and cleavage in Methylomicrobium alcaliphilum 20Z, a gene whose encoded sequence showed high similarity to sugar kinases of the ribokinase family was found. By heterologous expression of this gene in Escherichia coli cells and following metal chelate affinity chromatography, the electrophoretically homogenous recombinant enzyme with six histidine residues on the C-end was obtained. The enzyme catalyzes ATP-dependent phosphorylation of fructose into fructose-6-phosphate but is not active with other sugars as phosphoryl acceptors. The fructokinase of M. alcaliphilum 20Z is most active in the presence of Mn2+ at pH 9.0 and 60°C, being inhibited by ADP (K i = 2.50 ± 0.03 mM). The apparent K m values for fructose and ATP are 0.26 and 1.3 mM, respectively; the maximal activity is 141 U/mg protein. The enzyme shows the highest similarity of translated amino acid sequence with putative fructokinases of methylotrophic and autotrophic proteobacteria whose fruK gene is located in the gene cluster of sucrose biosynthesis. The involvement of fructokinase in sucrose metabolism in M. alcaliphilum 20Z and other methanotrophs and autotrophs is discussed.
Keywords: fructokinases; sucrose metabolism; methylotrophic bacteria; Methylomicrobium alcaliphilum ; fruK

DNA glycosylases play important roles in DNA repair in a variety of organisms, including humans. However, the function and regulation of these enzymes in the pathogenic bacterium Mycobacterium tuberculosis and related species are poorly understood. In the present study, the physical and functional interactions between 3-methyladenine DNA glycosylase (MAG) and topoisomerase I (TopA) in M. tuberculosis and M. smegmatis were characterized. MAG was found to inhibit the function of TopA in relaxing supercoiled DNA. In contrast, TopA stimulated the cleavage function of MAG on a damaged DNA substrate that contains hypoxanthine. The interaction between the two proteins was conserved between the two mycobacterial species. Several mutations in MAG that led to the loss of its interaction with and activity regulation of TopA were also characterized. The results of this study further elucidate glycosylase regulation in both M. smegmatis and M. tuberculosis.
Keywords: Mycobacterium tuberculosis ; DNA glycosylase; topoisomerase I; DNA repair

Metabolic dysfunction and relationship in human frontoparietal cortex in severe traumatic brain injury: Single-voxel 1H magnetic resonance spectroscopy study by N. A. Semenova; T. A. Akhadov; A. V. Petryaykin; S. S. Sidorin; A. V. Lukovenkov; S. D. Varfolomeev (388-394).
1H-magnetic resonance spectroscopy revealed that apparently normal (from the data of magnetic resonance imaging) human brain frontoparietal cortex in the subacute stage of traumatic brain injury is characterized by decreased level of N-acetylaspartate (NAA) and increase in levels of myoinositol, choline-containing compounds (Cho), and creatine/phosphocreatine (Cr). Correlations between Cr, Cho, and NAA were established. We propose a scheme of neuronal metabolic processes that joins these substances.
Keywords: magnetic resonance spectroscopy; severe traumatic brain injury; frontoparietal cortex

Glucuronoarabinoxylan is a key tethering glucan in the primary cell wall of cereals. Glucuronoarabinoxylan was extracted from different zones of maize (Zea mays L.) roots using endoxylanase that specifically cleaves β-(1,4)-glycoside bond between two consequent unsubstituted xylose residues. Changes in polysaccharide structure during elongation growth were characterized. Glucuronoarabinoxylan extractable after the endoxylanase treatment consisted of high molecular weight (30–400 kDa) and low molecular weight (<10 kDa) fractions. The presence of high molecular weight derivatives indicated that part of the natural glucuronoarabinoxylan is not digestible by the endoxylanase. This could be due to the revealed peculiar structural features, such as high level of substitution of xylose, absence of unsubstituted xylose residues existing in sequence, and significant degree of acetylation. In maize root meristem the indigestible fraction was 98% of the total extracted glucuronoarabinoxylan. This portion decreases to 47% during elongation. Also, the average molecular weight of indigestible glucuronoarabinoxylan reduced twofold. These changes in the ratio of glucuronoarabinoxylan fragments with different structure during root cell growth could reflect a transition of polysaccharide from its separating (highly substituted indigestible glucuronoarabinoxylan) form to that binding to cellulose microfibrils or other glucuronoarabinoxylan molecules and, hence, retarding growth.
Keywords: glucuronoarabinoxylan; endoxylanase; cell wall; elongation; growth; Zea mays

Nitrate reductase from Triticum aestivum leaves: Regulation of activity and possible role in production of nitric oxide by E. I. Galeeva; T. V. Trifonova; A. A. Ponomareva; L. V. Viktorova; F. V. Minibayeva (404-410).
Nitrate reductase (NR) and peroxidase (POX) are important enzymes involved in the metabolism of reactive oxygen (ROS) and nitrogen species in leaves of wheat (Triticum aestivum L.) seedlings. It has been confirmed that NR activity in wheat leaves depends on the light conditions and the presence of nitrates during the cultivation of the seedlings, and it is regulated by the molybdenum cofactor and phosphorylation. In the present study, confocal microscopy and EPR spectroscopy studies showed that the addition of nitrite, a product of NR, increased the level of nitric oxide (NO). This increase was prevented by the addition of sodium azide, an inhibitor of NR. The results suggest that in wheat leaves one of the key functions of NR is the formation of the signaling NO molecule. Cultivation of green plants under conditions of prolonged (4 days) darkness, a strong stress factor for photosynthesizing cells, decreased the activity of NR. Moreover, darkness induced significant elevation of the POX activity that was prevented by the addition of nitrate to the growth medium. It is proposed that the changes in light conditions result in the competition between nitrate- and ROS-metabolizing activities of POX in leaves, and a possible interaction between NR and POX controls the levels of NO and ROS in the leaf tissue.
Keywords: reactive oxygen species; nitrate reductase; nitric oxide; peroxidase; Triticum aestivum L.

In most insects and some other protostomes, a small stretch of nucleotides can be removed from mature 28S rRNA molecules, which could create two 28S rRNA subunits (28Sα and 28Sβ). Thus, during electrophoresis, the rRNA profiles of these organisms may differ significantly from the standard benchmark since the two subunits co-migrate with the 18S rRNA. To understand the structure and mechanism of the atypical 28S rRNA molecule, partial fragments of 28Sα and 28Sβ in brine shrimp Artemia parthenogenetica and planarian Dugesia japonica were cloned using a modified technology based on terminal transferase. Alignment with the corresponding sequences of 28S rDNAs indicates that there are 41 nucleotides in A. parthenogenetica and 42 nucleotides in D. japonica absent from the mature rRNAs. The AU content of the gap sequences of D. japonica and A. parthenogenetica is high. Both the gaps may form stem-loop structure. In D. japonica a UAAU cleavage signal is identified in the loop, but it is absent in A. parthenogenetica. Thus, it is proposed that the gap processing of 28S rRNA was a late enzyme-dependent cleavage event in the rRNA maturational process based on the AU rich gap sequence and the formation of the stem-loop structure to expose the processing segment, while the deletion of the gap region would not affect the structure and function of the 28S rRNA molecule.
Keywords: 28S rRNA; gap region; fragmentation; Artemia parthenogenetica ; Dugesia japonica