Biochemistry (Moscow) (v.80, #13)

Role of Small Noncoding RNAs in Bacterial Metabolism by T. L. Azhikina; D. V. Ignatov; E. G. Salina; M. V. Fursov; A. S. Kaprelyants (1633-1646).
The study of prokaryotic small RNAs is one of the most important directions in modern molecular biology. In the last decade, multiple short regulatory transcripts have been found in prokaryotes, and for some of them functional roles have been elucidated. Bacterial small RNAs are implicated in the regulation of transcription and translation, and they affect mRNA stability and gene expression via different mechanisms, including changes in mRNA conformation and interaction with proteins. Most small RNAs are expressed in response to external factors, and they help bacteria to adapt to changing environmental conditions. Bacterial infections of various origins remain a serious medical problem, despite significant progress in fighting them. Discovery of mechanisms that bacteria employ to survive in infected organisms and ways to block these mechanisms is promising for finding new treatments for bacterial infections. Regulation of pathogenesis with small RNAs is an attractive example of such mechanisms. This review considers the role of bacterial small RNAs in adaptation to stress conditions. We pay special attention to the role of small RNAs in Mycobacterium tuberculosis infection, in particular during establishment and maintenance of latent infection.
Keywords: bacteria; small noncoding RNAs; Hfq; regulation of gene expression; stress; virulence; Mycobacterium tuberculosis

Bacterial Small Regulatory RNAs and Hfq Protein by V. N. Murina; A. D. Nikulin (1647-1654).
Small regulatory RNA (sRNA) is a unique noncoding RNA involved in regulation of gene expression in both eukaryotic and bacterial cells. This short review discusses examples of positive and negative translation regulation by sRNAs in bacteria and participation of Hfq in these processes. The importance of structure investigation of nucleotide–protein and RNA–protein complexes for designing a model of Hfq interaction with both mRNA and sRNA simultaneously is demonstrated.
Keywords: Hfq; small regulatory RNA; sRNA; RNA–protein recognition; regulation of gene expression

Carbonyl Stress in Bacteria: Causes and Consequences by O. V. Kosmachevskaya; K. B. Shumaev; A. F. Topunov (1655-1671).
Pathways of synthesis of the α-reactive carbonyl compound methylglyoxal (MG) in prokaryotes are described in this review. Accumulation of MG leads to development of carbonyl stress. Some pathways of MG formation are similar for both pro- and eukaryotes, but there are reactions specific for prokaryotes, e.g. the methylglyoxal synthase reaction. This reaction and the glyoxalase system constitute an alternative pathway of glucose catabolism–the MG shunt not associated with the synthesis of ATP. In violation of the regulation of metabolism, the cell uses MG shunt as well as other glycolysis shunting pathways and futile cycles enabling stabilization of its energetic status. MG was first examined as a biologically active metabolic factor participating in the formation of phenotypic polymorphism and hyperpersistent potential of bacterial populations. The study of carbonyl stress is interesting for evolutionary biology and can be useful for constructing highly effective producer strains.
Keywords: carbonyl stress; bacteria; methylglyoxal; metabolite overproduction

Sperm-Specific Glyceraldehyde-3-Phosphate Dehydrogenase–An Evolutionary Acquisition of Mammals by V. I. Muronetz; M. L. Kuravsky; K. V. Barinova; E. V. Schmalhausen (1672-1689).
This review is focused on the mammalian sperm-specific glyceraldehyde-3-phosphate dehydrogenase (GAPDS). GAPDS plays the major role in the production of energy required for sperm cell movement and does not perform non-glycolytic functions that are characteristic of the somatic isoenzyme of glyceraldehyde-3-phosphate dehydrogenase. The GAPDS sequence is composed of 408 amino acid residues and includes an additional N-terminal region of 72 a.a. that binds the protein to the sperm tail cytoskeleton. GAPDS is present only in the sperm cells of mammals and lizards, possibly providing them with certain evolutionary advantages in reproduction. In this review, studies concerning the problems of GAPDS isolation, its catalytic properties, and its structural features are described in detail. GAPDS is much more stable compared to the somatic isoenzyme, perhaps due to the necessity of maintaining the enzyme function in the absence of protein expression. The site-directed mutagenesis approach revealed the two GAPDS-specific proline residues, as well as three salt bridges, which seem to be the basis of the increased stability of this protein. As distinct from the somatic isoenzyme, GAPDS exhibits positive cooperativity in binding of the coenzyme NAD+. The key role in transduction of structural changes induced by NAD+ is played by the salt bridge D311–H124. Disruption of this salt bridge cancels GAPDS cooperativity and twofold increases its enzymatic activity instead. The expression of GAPDS was detected in some melanoma cells as well. Its role in the development of certain pathologies, such as cancer and neurodegenerative diseases, is discussed.
Keywords: glyceraldehyde-3-phosphate dehydrogenase; sperm-specific glyceraldehyde-3-phosphate dehydrogenase; GAPDH; evolution of GAPDH; stability of GAPDH; sperm motility; glycolysis; melanoma cells; oncomarker; NAD-binding

Role of a Structurally Equivalent Phenylalanine Residue in Catalysis and Thermal Stability of Formate Dehydrogenases from Different Sources by V. I. Tishkov; K. V. Goncharenko; A. A. Alekseeva; S. Yu. Kleymenov; S. S. Savin (1690-1700).
Comparison of amino acid sequences of NAD+-dependent formate dehydrogenases (FDH, EC from different sources and analysis of structures of holo-forms of FDH from bacterium Pseudomonas sp. 101 (PseFDH) and soya Glycine max (SoyFDH) as well as of structure of apo-form of FDH from yeast Candida boidinii (CboFDH) revealed the presence on the surface of protein globule of hydrophobic Phe residue in structurally equivalent position (SEP). The residue is placed in the coenzyme-binding domain and protects bound NAD+ from solvent. The effects of amino acid changes of the SEP on catalytic properties and thermal stability of PseFDH, SoyFDH, and CboFDH were compared. The strongest effect was observed for SoyFDH. All eight amino acid replacements resulted in increase in thermal stability, and in seven cases, increase in catalytic constant was achieved. Thermal stability of SoyFDH after mutations Phe290Asp and Phe290Glu increased 66- and 55-fold, respectively. K M values of the enzyme for substrate and coenzyme in different cases slightly increased or decreased. In case of one CboFDH, the mutein catalytic constant increased and thermal stability did not changed. In case of the second CboFDH mutant, results were the opposite. In one PseFDH mutant, amino acid change did not influence the catalytic constant, but in three others, the parameter was reduced. Two PseFDH mutants had higher and two mutants lower thermal stability in comparison with initial enzyme. Analysis of results of SEP mutagenesis in FDHs from bacterium, yeast, and plant shows that protein structure plays a key role for effect of the same amino acid changes in equivalent position in protein globule of formate dehydrogenases from different sources.
Keywords: formate dehydrogenase; rational design; thermal stability; catalytic properties

Plant Proteases Involved in Regulated Cell Death by A. A. Zamyatnin Jr. (1701-1715).
Each plant genome encodes hundreds of proteolytic enzymes. These enzymes can be divided into five distinct classes: cysteine-, serine-, aspartic-, threonine-, and metalloproteinases. Despite the differences in their structural properties and activities, members of all of these classes in plants are involved in the processes of regulated cell death–a basic feature of eukaryotic organisms. Regulated cell death in plants is an indispensable mechanism supporting plant development, survival, stress responses, and defense against pathogens. This review summarizes recent advances in studies of plant proteolytic enzymes functioning in the initiation and execution of distinct types of regulated cell death.
Keywords: programmed cell death; PCD; apoptosis; autophagy; vacuolar processing enzyme; metacaspase; phytaspase; papain-like protease; proteasome

This review covers studies on the dependence of chlorophyll photobiosynthesis reactions from protochlorophyllide on the spectral composition of actinic light. A general scheme of the reaction sequence for the photochemical stage in chlorophyll biosynthesis for etiolated plant leaves is presented. Comparative analysis of the data shows that the use of light with varied wavelengths for etiolated plant illumination reveals parallel transformation pathways of different protochloro-phyllide forms into chlorophyllide, including a pathway for early photosystem II reaction center P-680 pigment formation.
Keywords: chlorophyll; protochlorophyllide; photoreduction; fluorescence spectra; absorption spectra

Regulation of Zygotic Genome and Cellular Pluripotency by D. V. Onichtchouk; A. S. Voronina (1723-1733).
Events, manifesting transition from maternal to zygotic period of development are studied for more than 100 years, but underlying mechanisms are not yet clear. We provide a brief historical overview of development of concepts and explain the specific terminology used in the field. We further discuss differences and similarities between the zygotic genome activation and in vitro reprogramming process. Finally, we envision the future research directions within the field, where biochemical methods will play increasingly important role.
Keywords: zebrafish; ZGA; MBT; pluripotency; Oct4; Nanog

Small Heat Shock Proteins and Distal Hereditary Neuropathies by V. V. Nefedova; L. K. Muranova; M. V. Sudnitsyna; A. S. Ryzhavskaya; N. B. Gusev (1734-1747).
Classification of small heat shock proteins (sHsp) is presented and processes regulated by sHsp are described. Symptoms of hereditary distal neuropathy are described and the genes whose mutations are associated with development of this congenital disease are listed. The literature data and our own results concerning physicochemical properties of HspB1 mutants associated with Charcot–Marie–Tooth disease are analyzed. Mutations of HspB1, associated with hereditary motor neuron disease, can be accompanied by change of the size of HspB1 oligomers, by decreased stability under unfavorable conditions, by changes in the interaction with protein partners, and as a rule by decrease of chaperone-like activity. The largest part of these mutations is accompanied by change of oligomer stability (that can be either increased or decreased) or by change of intermonomer interaction inside an oligomer. Data on point mutation of HspB3 associated with axonal neuropathy are presented. Data concerning point mutations of Lys141 of HspB8 and those associated with hereditary neuropathy and different forms of Charcot–Marie–Tooth disease are analyzed. It is supposed that point mutations of sHsp associated with distal neuropathies lead either to loss of function (for instance, decrease of chaperone-like activity) or to gain of harmful functions (for instance, increase of interaction with certain protein partners).
Keywords: small heat shock proteins; phosphorylation; chaperone-like activity; cytoskeleton; congenital diseases

Investigations of Molecular Mechanisms of Actin–Myosin Interactions in Cardiac Muscle by L. V. Nikitina; G. V. Kopylova; D. V. Shchepkin; S. R. Nabiev; S. Y. Bershitsky (1748-1763).
The functional characteristics of cardiac muscle depend on the composition of protein isoforms in the cardiomyocyte contractile machinery. In the ventricular myocardium of mammals, several isoforms of contractile and regulatory proteins are expressed–two isoforms of myosin (V1 and V3) and three isoforms of tropomyosin chains (α, β, and κ). Expression of protein isoforms depends on the animal species, its age and hormonal status, and this can change with pathologies of the myocardium. Mutations in these proteins can lead to cardiomyopathies. The functional significance of the protein isoform composition has been studied mainly on intact hearts or on isolated preparations of myocardium, which could not provide a clear comprehension of the role of each particular isoform. Present-day experimental techniques such as an optical trap and in vitro motility assay make it possible to investigate the phenomena of interactions of contractile and regulatory proteins on the molecular level, thus avoiding effects associated with properties of a whole muscle or muscle tissue. These methods enable free combining of the isoforms to test the molecular mechanisms of their participation in the actin–myosin interaction. Using the optical trap and the in vitro motility assay, we have studied functional characteristics of the cardiac myosin isoforms, molecular mechanisms of the calcium-dependent regulation of actin–myosin interaction, and the role of myosin and tropomyosin isoforms in the cooperativity mechanisms in myocardium. The knowledge of molecular mechanisms underlying myocardial contractility and its regulation is necessary for comprehension of cardiac muscle functioning, its disorders in pathologies, and for development of approaches for their correction.
Keywords: cardiac myosin isoforms; tropomyosin; actin–myosin interaction; Ca2+ regulation; optical trap; in vitro motility assay

Diversity of Potassium Channel Ligands: Focus on Scorpion Toxins by A. I. Kuzmenkov; E. V. Grishin; A. A. Vassilevski (1764-1799).
Potassium (K+) channels are a widespread superfamily of integral membrane proteins that mediate selective transport of K+ ions through the cell membrane. They have been found in all living organisms from bacteria to higher multicellular animals, including humans. Not surprisingly, K+ channels bind ligands of different nature, such as metal ions, low molecular mass compounds, venom-derived peptides, and antibodies. Functionally these substances can be K+ channel pore blockers or modulators. Representatives of the first group occlude the channel pore, like a cork in a bottle, while the second group of ligands alters the operation of channels without physically blocking the ion current. A rich source of K+ channel ligands is venom of different animals: snakes, sea anemones, cone snails, bees, spiders, and scorpions. More than a half of the known K+ channel ligands of polypeptide nature are scorpion toxins (KTx), all of which are pore blockers. These compounds have become an indispensable molecular tool for the study of K+ channel structure and function. A recent special interest is the possibility of toxin application as drugs to treat diseases involving K+ channels or related to their dysfunction (channelopathies).
Keywords: ion channel; potassium channel; ligand; pore blocker; modulator; venom; scorpion; toxin

Beta-Amyloid and Tau-Protein: Structure, Interaction, and Prion-Like Properties by O. G. Tatarnikova; M. A. Orlov; N. V. Bobkova (1800-1819).
During the last twenty years, molecular genetic investigations of Alzheimer’s disease (AD) have significantly broadened our knowledge of basic mechanisms of this disorder. However, still no unambiguous concept on the molecular bases of AD pathogenesis has been elaborated, which significantly impedes the development of AD therapy. In this review, we analyze issues concerning processes of generation of two proteins (β-amyloid peptide and Tau-protein) in the cell, which are believed to play the key role in AD genesis. Until recently, these agents were considered independently of each other, but in light of the latest studies, it becomes clear that it is necessary to study their interaction and combined effects. Studies of mechanisms of toxic action of these endogenous compounds, beginning from their interaction with known receptors of main neurotransmitters to specific peculiarities of functioning of signal intracellular pathways upon development of this pathology, open the way to development of new pharmaceutical substances directed concurrently on key mechanisms of interaction of toxic proteins inside the cell and on the pathways of their propagation in the extracellular space.
Keywords: Alzheimer’s disease; β-amyloid; Tau-protein; APP; presenilins; prion-like mechanism of AD

Detection of Intermolecular Interactions Based on Surface Plasmon Resonance Registration by D. V. Sotnikov; A. V. Zherdev; B. B. Dzantiev (1820-1832).
Methods for registration of intermolecular interactions based on the phenomenon of surface plasmon resonance (SPR) have become one of the most efficient tools to solve fundamental and applied problems of analytical biochemistry. Nevertheless, capabilities of these methods are often insufficient to detect low concentrations of analytes or to screen large numbers of objects. That is why considerable efforts are directed at enhancing the sensitivity and efficiency of SPR-based measurements. This review describes the basic principles of the detection of intermolecular interactions using this method, provides a comparison of various types of SPR detectors, and classifies modern approaches to enhance sensitivity and efficiency of measurements.
Keywords: surface plasmon resonance, registration of intermolecular interactions, enhancement of analytical signal, multiparametric assay