Biochemistry (Moscow) (v.74, #13)

Proteasome system of protein degradation and processing by A. V. Sorokin; E. R. Kim; L. P. Ovchinnikov (1411-1442).
In eukaryotic cells, degradation of most intracellular proteins is realized by proteasomes. The substrates for proteolysis are selected by the fact that the gate to the proteolytic chamber of the proteasome is usually closed, and only proteins carrying a special “label” can get into it. A polyubiquitin chain plays the role of the “label”: degradation affects proteins conjugated with a ubiquitin (Ub) chain that consists at minimum of four molecules. Upon entering the proteasome channel, the polypeptide chain of the protein unfolds and stretches along it, being hydrolyzed to short peptides. Ubiquitin per se does not get into the proteasome, but, after destruction of the “labeled” molecule, it is released and labels another molecule. This process has been named “Ub-dependent protein degradation”. In this review we systematize current data on the Ub-proteasome system, describe in detail proteasome structure, the ubiquitination system, and the classical ATP/Ub-dependent mechanism of protein degradation, as well as try to focus readers’ attention on the existence of alternative mechanisms of proteasomal degradation and processing of proteins. Data on damages of the proteasome system that lead to the development of different diseases are given separately.
Keywords: proteasome; Ub; ubiquitin; degradation; processing

Topogenesis of mitochondrial proteins includes their synthesis in cytosol and mitochondria, their translocation across the outer and inner membranes, sorting to various mitochondrial compartments, and assembly of different protein complexes. These complexes are involved in transport functions, electron transfer through the respiratory chain, generation of transmembrane electrochemical potential, oxidative phosphorylation of ADP into ATP, etc. To perform these functions, a special stringent control is required over formation of submitochondrial structures and the mitochondrion as a whole. Such control is expected to rigorously eliminate not only misfolded proteins but also incorrectly incorporated subunits and is realized in mitochondria by means of numerous proteases with different functions and localizations. In the case of more complicated protein formations, e.g. supercomplexes, the protein quality is assessed by their ability to realize the integral function of the respiratory chain and, thus, ensure the stability of the whole system. Considering supercomplexes of the mitochondrial respiratory chain, the present review clearly demonstrates that this control is realized by means of various (mainly vacuolar) proteases with different functions and localizations. The contemporary experimental data also confirm the author’s original idea that the general mechanism of assembly of subcellular structures is based on the “selection by performance criterion” and “stabilization by functioning”.
Keywords: biogenesis of mitochondria; protein quality control; respiratory complexes; mitochondrial proteases

Nicking endonucleases by L. A. Zheleznaya; G. S. Kachalova; R. I. Artyukh; A. K. Yunusova; T. A. Perevyazova; N. I. Matvienko (1457-1466).
Nicking endonucleases are a new type of enzymes. Like restriction endonucleases, they recognize short specific DNA sequence and cleave DNA at a fixed position relatively to the recognition sequence. However, unlike restriction endonucleases, nicking endonucleases cleave only one predetermined DNA strand. Until recently, nicking endonucleases were suggested to be naturally mutated restriction endonucleases which had lost their ability to dimerize and as a result the ability to cleave the second strand. We have shown that nicking endonucleases are one of the subunits of heterodimeric restriction endonucleases. Mechanisms used by various restriction endonucleases for double-stranded cleavage, designing of artificial nicking endonucleases on the basis of restriction endonucleases, and application of nicking endonucleases in molecular biology are reviewed.
Keywords: nicking endonuclease; restriction endonuclease; heterodimeric restriction endonuclease

Structure and mechanism of action of type IA DNA topoisomerases by D. V. Bugreev; G. A. Nevinsky (1467-1481).
DNA topoisomerases are enzymes responsible for regulation of genomic DNA supercoiling. They participate in essential processes of cells such as replication, transcription, recombination, repair, etc., and they are necessary for normal functioning of the cells. Topoisomerases alter the topological state of DNA by either passing one strand of the helix through the other strand (type I) or by passing a region of duplex DNA through another region of duplex DNA (type II). Type I DNA topoisomerases are subdivided into enzymes that bind to the 5′-(type IA) or 3′-phosphate group (type IB) during relaxation of the cleavable DNA. This review summarizes the literature on type IA DNA topoisomerases. Special attention is given to particular properties of their structure and mechanisms of functioning of these enzymes.
Keywords: type IA DNA topoisomerases; structure; reaction mechanism

From structure and functions of steroidogenic enzymes to new technologies of gene engineering by L. A. Novikova; Ya. V. Faletrov; I. E. Kovaleva; S. Mauersberger; V. N. Luzikov; V. M. Shkumatov (1482-1504).
This review summarizes data about structural and functional organization of steroidogenic P450-dependent enzymatic systems. Problems of catalysis of steroid substrate transformation, special features of mitochondrial type P450scc topogenesis, and abilities of some microbial electron transport proteins to support P450 activity in vitro and in vivo are considered. Principal steps in the creation and catalytic properties of transgenic strains of Escherichia coli, Saccharomyces cerevisiae, and Yarrowia lipolytica expressing both mammalian steroidogenic P450s and the corresponding electron transport proteins are also described. Achievements and prospects of using such transgenic strains for biotechnological synthesis and pharmacological screening are considered.
Keywords: cytochrome P450; steroid hormone biosynthesis; transgenic microorganisms; topogenesis; electron transport proteins; cytochrome P450 inhibitors

Molecular diversity of spider venom by A. A. Vassilevski; S. A. Kozlov; E. V. Grishin (1505-1534).
Spider venom, a factor that has played a decisive role in the evolution of one of the most successful groups of living organisms, is reviewed. Unique molecular diversity of venom components including substances of variable structure (from simple low molecular weight compounds to large multidomain proteins) with different functions is considered. Special attention is given to the structure, properties, and biosynthesis of toxins of polypeptide nature.
Keywords: spider venom; toxin; neurotoxin; cytolytic peptide; acylpolyamine; latrotoxin; toxic enzymes; protein structure and function; biosynthesis

The pathways of transformation of the chromophore of pigment-protein complexes have been studied at the terminal light-dependent stage of chlorophyll biosynthesis in plant leaves. The overall scheme of the sequence of photochemical and dark reactions of the pigment chromophore initiated by the reaction of photochemical hydration of a molecule of the precursor (protochlorophyllide) is presented. Schemes of the transformations of the components of the photoactive protochlorophyllide-oxidoreductase complex are discussed. Data are presented of features of the process at different stages of the formation of the pigment apparatus of plants.
Keywords: chlorophyll; chlorophyll biosynthesis; protochlorophyllide; protochlorophyllide-oxidoreductase; photosystems

Free radicals and cell chemiluminescence by Yu. A. Vladimirov; E. V. Proskurnina (1545-1566).
Application of chemiluminescence (CL) for study of free-radical reactions in human and animal cells and tissues is considered in this review. Historically, the study of intrinsic (ultraweak) luminescence gave place to the measurement of CL in the presence of chemical activators (CL probes) and physical activators (sensitizers) of luminescence, which made the method much more sensitive and specific. The methods of CL and EPR are direct methods of radical investigation, though the advantage of the CL method consists in the fact that CL intensity is directly proportional to a steady-state concentration of the radicals responsible for luminescence (first of all, lipid and oxygen radicals) irrespective the activity of these radicals. The mechanisms of CL reactions in the absence of activators and in the presence of luminol and lucigenin are considered. Examples of various applications of the CL method in medical, biological, and clinical investigations are given including those for estimation of the phagocytic activity of cells, antioxidant activity, determination of toxicity, and other purposes.
Keywords: chemiluminescence; reactive oxygen species; luminol; lucigenin; cell luminescence; lipid peroxidation

This review deals with artificial modular nanotransporters (MNT) of polypeptide nature for drug delivery into target cells and then into a specified cell compartment like the nucleus. The developed approach is based on the use of intra-cellular transport processes characteristic of practically all cells, including cancer cells. The first MNT module ligand carries out a double function: specific recognition of a cancer target cell and penetration into the cell via receptor-mediated endocytosis. The movement of the MNT within the cell along this path specifies the need to supply the MNT with an endosomolytic module making it possible to leave the endocytotic pathway before getting into lysosomes in order to have time for interaction with importins. For this purpose, a polypeptide fragment able to make defects in membranes only at the pH of endosomes is used as the second module. Delivery into the cell nucleus is provided by the third module containing an amino acid sequence of nuclear localization, “recognized” by importins located in the hyaloplasm. And finally, the fourth module, a carrier for joining the transported drug, is incorporated into the MNT. Depending on the type of ligand module, MNT for different target cell types have been produced. Each module retains its activity within the MNT, ligand modules bind target receptors with high affinity, while the module with the nuclear localization sequence binds importins. The endosomolytic module forms pores in lipid membranes through which MNT are able to leave acidifying cell compartments (endosomes). Modules within MNT can be replaced or transposed, which makes it possible to use them for delivery of different drugs into different target cells and their compartments. It was shown that photosensitizers and radionuclides used for cancer therapy acquire pronounced cell specificity as well as the 10-1000-fold higher efficiency resulting from their delivery into the most vulnerable compartment — the cell nucleus.
Keywords: modular drug nanotransporters; drug delivery; photosensitizers; alpha-emitters; ErbB1 over-expression; intra-cellular transport; cancer therapy

Fragmentomics of natural peptide structures by A. A. Zamyatnin (1575-1585).
Natural fragmentation of peptide and other chemical structures is well known. They are a significant object of biochemical investigations. In this connection, the bases and determination are given for the notion of the “fragmentome” as a set of all fragments of a single substance, as well as for global fragmentome of all chemical components of living organisms. It is described how protein-peptide fragments are formed in nature, what experimental and theoretical methods are used for their investigation, as well as mathematical characteristics of fragmentomes. Individual fragmentomes of all subunits and of complete casein fragmentome are considered in detail. Structural and functional variety of its possible fragments was revealed by computer analysis. Formation in an organism of an exogenous-endogenous pool of oligopeptides and correlation of these data with concepts of structure-functional continuum of regulatory molecules is shown on an example of food protein fragments. Possible practical importance of the use of natural fragments in dietology, therapy, as well as in sanitary hygiene and cosmetics is noted.
Keywords: fragmentomics; oligopeptide; protein; casein; structure; function; EROP-Moscow Database

Dynamic proteomics in modeling of the living cell. Protein-protein interactions by A. A. Terentiev; N. T. Moldogazieva; K. V. Shaitan (1586-1607).
This review is devoted to describing, summarizing, and analyzing of dynamic proteomics data obtained over the last few years and concerning the role of protein-protein interactions in modeling of the living cell. Principles of modern high-throughput experimental methods for investigation of protein-protein interactions are described. Systems biology approaches based on integrative view on cellular processes are used to analyze organization of protein interaction networks. It is proposed that finding of some proteins in different protein complexes can be explained by their multi-modular and polyfunctional properties; the different protein modules can be located in the nodes of protein interaction networks. Mathematical and computational approaches to modeling of the living cell with emphasis on molecular dynamics simulation are provided. The role of the network analysis in fundamental medicine is also briefly reviewed.
Keywords: systems biology; modeling of living cells; dynamic proteomics; interactomes; protein interaction networks; structural and functional mapping of proteins; molecular dynamics; fundamental medicine