Biochemistry (Moscow) (v.76, #3)

Translation initiation factor 2 (IF2) is one of key components of the translation initiation system in living cells. In bacteria IF2 is a multidomain monomeric protein, while in eukaryotic and archaean cells e/aIF2 is heterotrimer (αβγ). Data, including our own, on eukaryotic type translation initiation factor 2 (e/aIF2) structure and functioning are presented. There are also new data on initiation factors eIF5 and eIF2B that directly interact with eIF2 and control its participation in nucleotide exchange.
Keywords: translation initiation; e/aIF2; heterotrimer; initiator methionyl-tRNA (Met-tRNAi); Met-tRNAi·e/aIF2·GTP triple complex

IgG-binding proteins of bacteria by E. V. Sidorin; T. F. Solov’eva (295-308).
Proteins capable of non-immune binding of immunoglobulins G (IgG) of various mammalian species, i.e. without the involvement of the antigen-binding sites of the immunoglobulins, are widespread in bacteria. These proteins are located on the surface of bacterial cells and help them to evade the host’s immune response due to protection against the action of complement and to decrease in phagocytosis. This review summarizes data on the structure of immunoglobulin-binding proteins (IBP) and their complexes with IgG. Common and distinctive structural features of IBPs of gram-positive bacteria (staphylococci, streptococci, peptostreptococci) are discussed. Conditions for IBP expression by bacteria and their functional heterogeneity are considered. Data on IBPs of gram-negative bacteria are presented.
Keywords: immunoglobulin G; immunoglobulin-binding proteins; Fc-fragment of IgG

Endogenous opioid peptides comprise a group of bioregulatory factors involved in regulation of functional activity of various physiological systems of an organism. One of most important functions of endogenous opioids is their involvement in the interaction between cells of the nervous and immune systems. Summary data on the effects of opioid peptides on regulation of functions of innate immunity cells are presented.
Keywords: opioid peptides; opiate receptors; immunoregulatory effects; monocytes/macrophages; neutrophils; NK cells; dendritic cells; cAMP

Plant cortical microtubules are putative sensors under abiotic stresses by Che Wang; Lijun Zhang; Wenfu Chen (320-326).
In this article, we review current knowledge on the dynamic changes and roles of microtubule (MT) arrays under abiotic stresses. The results emphasize the existence of highly dynamic changes, complex regulatory networks, and the vitally important role of MTs in the response to abiotic stresses. In particular, some findings indicate that cortical microtubules (CMTs) underlying the plasma membrane play an important role in abiotic stress-induced signaling pathways. Therefore, we also discuss the relationship between CMTs and abiotic stress signaling. The data show that at least three early response mechanisms, namely, Ca2+ signaling, abscisic acid biosynthesis, and the formation of plant cell walls, follow CMT reorganization and are mediated by dynamic changes in the CMTs. Consequently, we propose that the CMTs are not only part of the plant response to abiotic stresses but might also serve as a type of cell wall membrane-bound sensor that perceives the stress stimuli to generate adaptive signals and responses of cells.
Keywords: microtubules; abiotic stresses; response mechanisms; signaling pathway; sensors

Mucoadhesive polyelectrolyte microparticles containing recombinant human insulin and its analogs aspart and lispro by N. G. Balabushevich; M. A. Pechenkin; I. N. Zorov; E. D. Shibanova; N. I. Larionova (327-331).
Microparticles containing recombinant human insulin and its analogs aspart and lispro were prepared using an alternate adsorption of chitosan and dextran sulfate from solutions onto microaggregates of protein-dextran sulfate insoluble complex. The following properties of polyelectrolyte hormone-containing microparticles were studied: pH stability, surface charge, mucoadhesive properties, Ca2+ binding, degradation under the influence of proteases (trypsin, chymotrypsin). The influence of the self-association ability of encapsulated insulins on the form of protein releasing from microparticles was studied. Insulins aspart and lispro released from the microparticles as monomers were more liable to proteolysis than human insulin released as a hexamer. The combined effect of properties of polyelectrolyte microparticles and of encapsulated recombinant proteins on the bioavailability of insulin under peroral administration is discussed.
Keywords: polyelectrolyte microparticles; human insulin; insulin lispro and aspart; mucoadhesive properties; Ca2+ binding; proteolysis

Amino acid sequences of two immune-dominant epitopes of recoverin are involved in Ca2+/recoverin-dependent inhibition of phosphorylation of rhodopsin by I. I. Senin; N. K. Tikhomirova; V. A. Churumova; I. I. Grigoriev; T. A. Kolpakova; D. V. Zinchenko; P. P. Philippov; E. Yu. Zernii (332-338).
Antibodies AB60–72 and AB80–92 against two immune-dominant epitopes of photoreceptor Ca2+-binding protein recoverin, 60-DPKAYAQHVFRSF-72 and 80-LDFKEYVIALHMT-92, which can be exposed in a Ca2+-dependent manner, were obtained. The presence of AB60–72 or AB80–92 results in a slight increase in Ca2+-affinity of recoverin and does not affect significantly a Ca2+-myristoyl switch mechanism of the protein. However in the presence of AB60–72 or AB80–92 recoverin loses its ability to interact with rhodopsin kinase and consequently to perform a function of Ca2+-sensitive inhibitor of rhodopsin phosphorylation in photoreceptor cells.
Keywords: Ca2+-dependent antibodies; Ca2+-myristoyl switch; cancer associated retinopathy; recoverin

Foot and mouth disease virus polyepitope protein produced in bacteria and plants induces protective immunity in guinea pigs by E. P. Andrianova; S. R. Krementsugskaia; N. N. Lugovskaia; T. K. Mayorova; V. V. Borisov; M. A. Eldarov; N. V. Ravin; A. S. Folimonov; K. G. Skryabin (339-346).
The goal of this project was to develop an alternative foot and mouth disease (FMD) vaccine candidate based on a recombinant protein consisting of efficient viral epitopes. A recombinant gene was designed that encodes B-cell epitopes of proteins VP1 and VP4 and T-cell epitopes of proteins 2C and 3D. The polyepitope protein (H-PE) was produced in E. coli bacteria or in N. benthamiana plants using a phytovirus expression system. The methods of extraction and purification of H-PE proteins from bacteria and plants were developed. Immunization of guinea pigs with the purified H-PE proteins induced an efficient immune response against foot and mouth disease virus (FMDV) serotype O/Taiwan/99 and protection against the disease. The polyepitope protein H-PE can be used as a basis for developing a new recombinant vaccine against FMD.
Keywords: FMDV; vaccine; transient expression in plants; polyepitope protein

A gene encoding the xylanase from Bacillus subtilis strain R5 containing the native signal sequence was cloned and expressed in Escherichia coli. The heterologous expression of the gene resulted in the production of the recombinant protein in the cytoplasm as well as its secretion into the culture medium. The xylanase activity in the culture medium increased with time after induction up to 90% of the total activity in 14 h. Molecular mass and N-terminal amino acid sequence determinations of the purified recombinant xylanase revealed that the native signal peptide was cleaved off by E. coli signal peptidases between Ala28 and Ala29.
Keywords: xylanase; Bacillus subtilis strain R5; signal peptide; purification; MALDI-TOF mass spectrometry

Most mycobacteria appear to be naturally resistant to β-lactam antibiotics such as penicillin. However, very few β-lactamases and their regulation have been clearly characterized in Mycobacterium tuberculosis H37Rv. In this study, a unique bifunctional protein, Rv2752c, from M. tuberculosis showed both β-lactamase and RNase activities. Two residues, D184 and H397, appear to be involved in Zn2+-binding and are essential for the dual functions. Both activities are lost upon deletion of the C-terminal 100 a.a. long Rv2752c tail, which contains an additional loop when compared with the RNase J of Bacillus subtilis. A chaperone-like protein, Rv2373c, physically interacted with Rv2752c and inhibited both activities. This is the first report of characterization of a bifunctional β-lactamase and its regulation in mycobacteria. These data offered important clues for further investigation of the structure and function of microbial β-lactamases. Increased understanding of this protein will provide further insights into the mechanism of microbial drug resistance.
Keywords: Mycobacterium tuberculosis ; metallo-β-lactamase; ribonuclease

Major chondroitin sulfate proteoglycans identified in L6J1 myoblast culture by I. I. Ermakova; G. A. Sakuta; M. A. Potekhina; M. A. Fedorova; R. Hoffmann; V. I. Morozov (359-365).
The major proteoglycans from L6J1 rat myoblast culture were identified. The proteoglycans were isolated from different constituents of cell culture: culture medium, extracellular matrix (ECM), and myoblasts. To identify their core proteins, the proteoglycans were treated with enzymes specifically digesting chondroitin/dermatan sulfates or chondroitin sulfates. Subsequent electrophoresis and mass spectrometry revealed versican, collagen XII, and inter-α-trypsin inhibitor classified as chondroitin sulfate proteoglycans and biglycan known to be chondroitin/dermatan sulfate proteoglycan. Versican was identified in ECM and the other proteoglycans in the culture medium. Such difference in localization is likely to be a consequence of different biological functions. Versican, collagen XII, and biglycan are synthesized by myoblasts and inter-α-trypsin inhibitor originates from fetal bovine serum (a culture medium component).
Keywords: proteoglycans; glycosaminoglycans; myoblasts

Modeling amyloid fibril formation by N. V. Dovidchenko; O. V. Galzitskaya (366-373).
No detailed step-by-step model of protein rearrangements during amyloid structure formation has been presented in the literature. The aim of this work was to design a kinetic model for description of the amyloid formation process on the basis of the most recent experimental data. A general kinetic model is proposed for description of the amyloid formation process including the nucleation mechanism of polymerization with consecutive monomer attachment to oligomer and auto-catalytic growth of amyloid aggregates implying all types of exponential growth such as branching, fragmentation, and growth from the surface. Computer simulations have shown that the model correctly describes experimentally observed growth stages of amyloid fibrils and that the presence of exponential growth stage in the model is critical for modeling amyloid fibril formation. The key feature of the proposed model is the stage of the exponential growth of the aggregate. Such stage can simultaneously describe several versions of aggregate enlargement by branching, fragmentation, or growth from the surface. Data obtained using this model suggest conclusions concerning the significance of each stage in amyloid fibril assembly.
Keywords: amyloid fibril; critical concentration; lag period; aggregation; oligomeric particle

Britton Chance by V. P. Skulachev (374-375).