Biochemistry (Moscow) (v.76, #12)

Mechanisms of autophagy and pexophagy in yeasts by A. A. Sibirny (1279-1290).
Autophagy is a process of recycling of the intracellular constituents using vacuoles (lysosomes). General autophagy occurs due to involvement of highly conservative components found in all eukaryotes, from yeasts to higher plants and humans. Autophagy also could be a selective process and be involved in regulation of the cellular number of organelles, including that of peroxisomes. The process of specific autophagic peroxisome degradation is known as pexophagy. Yeasts appear to be convenient model for studying molecular mechanisms of pexophagy, and most known ATG genes (from the term AuTophaGy) were identified in yeast studies. This review examines characteristics of general autophagy, other types of autophagy as well as pexophagy, in particular, functions of Atg proteins in general autophagy and in macro- and micropexophagy. Special attention is given to mechanisms of phagophore assembly, the role of phosphatidylinositol-3-phosphate in pexophagy, the role of peroxines (proteins involved in peroxisome biogenesis) in pexophagy, as well as properties of Atg proteins specifically involved in micropexophagy.
Keywords: autophagy; pexophagy; macropexophagy; micropexophagy; Cvt pathway; Atg proteins; yeasts

Membrane lipids—phospholipids, fatty acids, and cholesterol—participate in thermal adaptation of ectotherms (bacteria, amphibians, reptiles, fishes) mainly via changes in membrane viscosity caused by the degree of fatty acids unsaturation, cholesterol/phospholipids ratio, and phospholipid composition. Studies of thermal adaptation of endotherms (mammals and birds) revealed the regulatory role of lipids in hibernation. Cholesterol and fatty acids participate in regulation of the parameters of torpor, gene expression, and activity of enzymes of lipid metabolism. Some changes in lipid metabolism during artificial and natural hypobiosis, namely, increased concentration of cholesterol and fatty acids in blood and decreased cholesterol concentration in neocortex, are analogous to those observed under stress conditions and coincide with mammalian nonspecific reactions to environmental agents. It is shown that the effects of artificial and natural hypobiosis on lipid composition of mammalian cell membranes are different. Changes in lipid composition cause changes in membrane morphology during mammalian hibernation. The effect of hypobiosis on lipid composition of membranes and cell organelles is specific and seems to be defined by the role of lipids in signaling systems. Comparative study of lipid metabolism in membranes and organelles during natural and artificial hypobiosis is promising for elucidation of adaptation of mammals to low ambient temperatures.
Keywords: mammals; hibernation; artificial hypobiosis; membranes; lipids

Diversity of integrase-hydrolyzing IgGs and IgMs from sera of HIV-infected patients by S. V. Baranova; V. N. Buneva; M. A. Kharitonova; L. P. Sizyakina; O. D. Zakharova; G. A. Nevinsky (1300-1311).
It was previously shown that small fractions of IgGs and IgMs from the sera of AIDS patients specifically hydrolyze only HIV integrase (IN) but not many other tested proteins. Here we present evidence showing that these IgGs and IgMs are extreme catalytically heterogeneous. Affinity chromatography on IN-Sepharose using elution of IgGs (or IgMs) with different concentration of NaCl and acidic buffer separated catalytic antibodies (ABs) into many AB subfractions demonstrating different values of K m for IN and k cat. Nonfractionated IgGs and IgMs possess serine-, thiol-, acidiclike, and metal-dependent proteolytic activity. Metal-dependent activity of abzymes increases in the presence of ions of different metals. In contrast to canonical proteases having one pH optimum, initial nonfractionated IgGs and IgMs demonstrate several optima at pH from 3 to 10. The data obtained show that IN-hydrolyzing polyclonal IgG and IgM of HIV-infected patients are cocktails of anti-IN ABs with different structure of the active centers possessing various affinity to IN, pH optima, and relative rates of the specific substrate hydrolysis.
Keywords: human immunodeficiency virus; abzymes; HIV infection; HIV-integrase

Changes in isoform composition, structure, and functional properties of titin from mongolian gerbil (Meriones unguiculatus) cardiac muscle after space flight by I. M. Vikhlyantsev; A. D. Okuneva; M. D. Shpagina; Yu. V. Shumilina; N. V. Molochkov; N. N. Salmov; Z. A. Podlubnaya (1312-1320).
Changes in isoform composition, secondary structure, and titin phosphorylation in Mongolian gerbil (Meriones unguiculatus) cardiac muscle were studied after 12-day-long space flight onboard the Russian spacecraft Foton-M3. The effect of titin on the actin-activated myosin ATPase activity at pCa 7.5 and 4.6 was also studied. Almost twofold increase in titin long N2BA isoform content relative to that of short N2B isoform was found on electrophoregrams of cardiac muscle left ventricle of the flight group gerbils. Differences in secondary structure of titin isolated from cardiac muscle of control and flight groups of gerbils were found. An increase in phosphorylation (1.30–1.35-fold) of titin of cardiac muscle of the flight group gerbils was found. A decrease in activating effect of titin of cardiac muscle of the flight group gerbils on actomyosin ATPase activity in vitro was also found. The observed changes are discussed in the context of M. unguiculatus cardiac muscle adaptation to conditions of weightlessness.
Keywords: Mongolian gerbils Meriones unguiculatus ; cardiac muscle; weightlessness; titin isoforms; actin-activated ATPase activity of myosin

Influence of intramolecular interactions on conformational and dynamic properties of analogs of heptapeptide AFP14–20 by N. T. Moldogazieva; K. V. Shaitan; M. Yu. Antonov; I. K. Vinogradova; A. A. Terentiev (1321-1336).
Conformational and dynamic properties of proteins and peptides play an important role in their functioning. However, mechanisms that underlie this influence have not been fully elucidated. In the present work we computationally constructed analogs of heptapeptide AFP14–20 (LDSYQCT) — one of the biologically active sites of human α-fetoprotein (AFP) — to study their conformational and dynamic properties using molecular dynamics simulation. Analogs were obtained by point substitutions of amino acid residues taking into account differences in their physicochemical properties and also on the basis of analysis of amino acid substitutions in the AFP14–20-like motifs revealed in different physiologically active proteins. It is shown that changes in conformational mobility of amino acid residues of analogs are due to disruption or arising of intramolecular interactions that, in turn, determine existence of steric restrictions during rotation around covalent bonds of the peptide backbone. Substitution of an amino acid by another one with significant difference in physicochemical properties may not lead to remarkable changes in conformational and dynamic properties of the peptide if intramolecular interactions remain unchanged.
Keywords: α-fetoprotein; AFP; analogs of heptapeptide AFP14–20 ; molecular dynamics; intramolecular interactions

Interaction of synthetic peptide octarphin with rat myocardium membranes by Y. N. Nekrasova; Y. A. Zolotarev; E. V. Navolotskaya (1337-1341).
A selective agonist of non-opioid β-endorphin receptor synthetic peptide octarphin (TPLVTLFK, specific activity 28 Ci/mmol) was prepared. The [3H]octarphin binding to rat myocardium membranes before and after experimental myocardial infarction (EMI) was studied. It was found that [3H]octarphin with high affinity and specificity binds to non-opioid β-endorphin receptor of rat myocardium membranes before EMI: K d1 value of the [3H]octarphin specific binding to membranes was 1.8 ± 0.2 nM. In 3 h after EMI a sharp lowering in affinity of the binding is observed (K d2 = 13.3 ± 0.4 nM), and in 48 h its almost complete restoration (K d4 = 2.2 ± 0.3 nM). The results indicate participation of non-opioid β-endorphin receptor in the regulation of myocardial activity.
Keywords: β-endorphin; naloxone; peptide; receptor; myocardium

Acetylation degree of chitin in the protective response of wheat plants by I. V. Maksimov; A. Sh. Valeev; R. F. Safin (1342-1346).
Influences on the acetylation degree of chitin manifested by proteins from cultural filtrates of strains of the fungus Septoria nodorum different in aggressiveness and of extracts from leaves of the susceptible (Triticum aestivum) and resistant (Triticum timopheevii) wheat plants infected with these strains were studied. Chitin deacetylase was found among the extracellular proteins of the fungus. Its activity was higher in the aggressive strain of the fungus than in the non-aggressive one, and this suggested that this enzyme could play an important role in the further formation of compatible relationship of the pathogens with the plants. Protein extracts from the susceptible wheat seedlings infected with the septoriosis agent also contained a component decreasing the acetylation degree of chitin. Protein extracts from the resistant wheat seedlings increased the chitin acetylation degree. It is supposed that this can be a pattern of the plant counteracting the action of chitin deacetylases of the pathogen.
Keywords: wheat (Triticum aestivum, Triticum timopheevii); septoriosis agent (Septoria nodorum); chitin; chitosan; acetylation degree

Classification and characterization of putative cytochrome P450 genes from Panax ginseng C. A. Meyer by Balusamy Sri Renuka Devi; Yu-Jin Kim; Subramaniyum Sathiyamoorthy; Altanzul Khorolragchaa; Sathiyaraj Gayathri; Shohana Parvin; Dong-Uk Yang; Senthil Kalai Selvi; Ok Ran Lee; Sungyoung Lee; Deok-Chun Yang (1347-1359).
In plants heme containing cytochrome P450 (P450) is a superfamily of monooxygenases that catalyze the addition of one oxygen atom from O2 into a substrate, with a substantial reduction of the other atom to water. The function of P450 families is attributed to chemical defense mechanism under terrestrial environmental conditions; several are involved in secondary and hormone metabolism. However, the evolutionary relationships of P450 genes in Panax ginseng remain largely unknown. In the present study, data mining methods were implemented and 116 novel putative P450 genes were identified from Expressed Sequence Tags (ESTs) of a ginseng database. These genes were classified into four clans and 22 families by sequence similarity conducted at amino acid level. The representative putative P450 sequences of P. ginseng and known P450 family from other plants were used to construct a phylogenetic tree. By comparing with other genomes, we found that most of the P450 genes from P. ginseng can be found in other dicot species. Depending on P450 family functions, seven P450 genes were selected, and for that organ specific expression, abiotic, and biotic studies were performed by quantitative reverse transcriptase-polymerase chain reaction. Different genes were found to be expressed differently in different organs. Biotic stress and abiotic stress transcript level was regulated diversely, and upregulation of P450 genes indicated the involvement of certain genes under stress conditions. The upregulation of the P450 genes under methyl jasmonate and fungal stress justifies the involvement of specific genes in secondary metabolite biosynthesis. Our results provide a foundation for further elucidating the actual function and role of P450 involved in various biochemical pathways in P. ginseng.
Keywords: abiotic; biotic; ESTs; gene expression; in silico ; Panax ginseng ; secondary metabolites

The change in the dark reduction rate of photooxidized reaction centers (RC) of type II from three anoxygenic bacteria (Rhodobacter sphaeroides R-26, Chromatium minutissimum, and Chloroflexus aurantiacus) having different redox potentials of the P+/P pair and availability of RC for exogenous electron donors was investigated upon the addition of Mn2+ and HCO 3 . It was found that the dark reduction of P 870 + from Rb. sphaeroides R-26 is considerably accelerated upon the combined addition of 0.5 mM MnCl2 and 30–75 mM NaHCO3 (as a result of formation of “low-potential” complexes [Mn(HCO3)2]), while MnCl2 and NaHCO3 added separately had no such effect. The effect is not observed either in RC from Cf. aurantiacus (probably due to the low oxidation potential of the primary electron donor, P865, which results in thermodynamic difficulties of the redox interaction between P 865 + and Mn2+) or in RC from Ch. minutissimum (apparently due to the presence of the RC-bound cytochrome preventing the direct interaction between P 870 + and Mn2+). The absence of acceleration of the dark reduction of P 870 + in the RC of Rb. sphaeroides R-26 when Mn2+ and HCO 3 were replaced by Mg2+ or Ca2+ and by formate, oxalate, or acetate, respectively, reveals the specificity of the Mn2+-bicarbonate complexes for the redox interaction with P+. The results of this work might be considered as experimental evidence for the hypothesis of the participation of Mn2+ complexes in the evolutionary origin of the inorganic core of the water oxidizing complex of photosystem II.
Keywords: Mn-bicarbonate complexes; reaction centers; purple bacteria; RC-bound cytochrome

In vivo injected mitochondria-targeted plastoquinone antioxidant SkQR1 prevents β-amyloid-induced decay of long-term potentiation in rat hippocampal slices by N. A. Kapay; N. K. Isaev; E. V. Stelmashook; O. V. Popova; D. B. Zorov; V. G. Skrebitsky; V. P. Skulachev (1367-1370).
Addition of 200 nM β-amyloid 1–42 (Abeta) to a rat hippocampal slice impairs the induction of a long-term post-tetanic potentiation (LTP) of population spike (PS) in pyramidal neurons of the CA1 field of hippocampus. Intraperitoneal injection into the rat of the mitochondria-targeted plastoquinone derivative SkQR1 (1 μmol/kg of weight given 24 h before the slices were made) abolishes the deleterious effect of Abeta on LTP. These data demonstrate that SkQR1 therapy is able to compensate the Abeta-induced impairments of long-term synaptic plasticity in the hippocampus, which are the main cause of loss of memory and other cognitive functions associated with Alzheimer’s disease.
Keywords: Alzheimer’s disease; β-amyloid; long-term memory; long-term potentiation; neurons; hippocampus; mitochondria-targeted antioxidants; SkQR1

Cell-penetrating peptides. Methods and protocols by G. Ya. Wiederschain (1371-1371).