T. F. Solov’eva

Interaction of chitosans and their N-acylated derivatives with lipopolysaccharide of gram-negative bacteria

The interactions of lipopolysaccharide (LPS) with the natural polycation chitosan and its derivatives—high molecular weight chitosans (80 kD) with different degree of acetylation, low molecular weight chitosan (15 kD), acylated oligochitosan (5.5 kD) and chitooligosaccharides (biose, triose, and tetraose)—were studied using ligand-enzyme solid-phase assay. The LPS-binding activity of chitosans (80 kD) decreased with increase in acetylation degree. Affinity of LPS interaction with chitosans increased after introduction of a fatty acid residue at the reducing end of chitosan. Activity of N-monoacylated chitooligosaccharides decreased in the order: oligochitosan → tetra-→ tri-→ disaccharides. The three-dimensional structures of complexes of R-LPS and chitosans with different degree of acetylation, chitooligosaccharides, and their N-monoacylated derivatives were generated by molecular modeling. The number of bonds stabilizing the complexes and the energy of LPS binding with chitosans decreased with increase in acetate group content in chitosans and resulted in changing of binding sites. It was shown that binding sites of chitooligosaccharides on R-LPS overlapped and chitooligosaccharide binding energies increased with increase in number of monosaccharide residues in chitosan molecules. The input of the hydrophobic fragment in complex formation energy is most prominent for complexes in water phase and is due to the hydrophobic interaction of chitooligosaccharide acyl fragment with fatty acid residues of LPS.

Chitosan antiviral activity: Dependence on structure and depolymerization method

Enzymatic (the action of lysozyme) and chemical (the action of hydrogen peroxide) hydrolysis of chitosans with various degree of acetylation (DA)—25, 17, and 1.5%—was performed. Purification and fractioning of the hydrolysis products were performed using dialysis, ultrafiltration, and gel-penetrating chromatography. Low-molecular (LM) derivatives of the polysaccharide with molecular masses from 17 to 2 kDa were obtained. The study of their antiviral activity against the tobacco mosaic virus (TMV) showed that these samples inhibited the formation of local necroses induced by the virus for 50–90%. The antiviral activity of the LM chitosans significantly increased with the lowering of their polymerization degree. Furthermore, the products of the enzymatic hydrolysis possessed lower activity than the chitosan samples obtained as a result of chemical hydrolysis. It was revealed that the exhibition of the antiviral activity weakly depended on the degree of acetylation of the samples.

Structure and properties of carrageenan-like polysaccharide from the red alga Tichocarpus crinitus (Gmel.) Rupr. (Rhodophyta, Tichocarpaceae)

Sulfated polysaccharides occurring in the red algae Tichocarpus crinitus cell wall were fractionated and purified. NMR and FT-IR spectroscopy analyses revealed that the non-gelling fraction contained a sulfated galactans having a new carrageenan-like structure. It is built with alternatively linked 1,3-linked β-D-galactopyranosyl-2,4-disulphates and 1,4-linked 3,6-anhydro-α-D-galactopyranosyl residues. Minor amounts of its biosynthetic precursor were detected in a water-extracted specimen. Brief analysis of rheological and biological properties of the non-gelling fraction was carried out. The carrageenan-like polysaccharide from T. crinitus displayed the properties of “random coil” polymer at high temperature, and possesses high anticoagulant activity at low concentration.

IgG-binding proteins of bacteria

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.

Influence of lipopolysaccharides and lipids A from some marine bacteria on spontaneous and Escherichia coli LPS-induced TNF-α release from peripheral human blood cells

Some endotoxic properties of lipopolysaccharides (LPS) and lipids A (LA) from the marine bacteria Marinomonas communis ATCC 27118T, Marinomonas mediterranea ATCC 700492T, and Chryseobacterium indoltheticum CIP 103168T were studied. The preparations tested were shown to have high 50% lethal doses (4 μg per mouse for LPS from M. mediterranea and more than 12 μg per mouse for two other LPS and LA from C. indoltheticum) and were moderate (371 ± 37 pg/ml at 10 μg/ml of C. indoltheticum LPS), weak (148 ± 5 pg/ml at 1 μg/ml of M. mediterranea LPS), and zero (LA and LPS from M. communis and LA from C. indoltheticum) inducers of tumor necrosis factor α (TNF-α) release from peripheral human blood cells. The capacity of the LA and LPS samples from marine bacteria to inhibit TNF-α release induced by LPS from Escherichia coli O55: B5 (10 ng/ml) was also studied.

In Vitro and Ex Vivo Studies of Antioxidant Activity of Carrageenans, Sulfated Polysaccharides from Red Algae

Antioxidant properties of structurally different sulfated polysaccharides (carrageenans) were studied in vitro and ex vivo. Ferric reducing antioxidant activity of carrageenans and their inhibitory effects on hydroxyl radicals and superoxide anion radicals were demonstrated in vitro. Activity of carrageenans depends on the polysaccharide structure. Carrageenans stimulate catalytic activity of SOD from donor erythrocyte.

Isolation and characterization of a low-molecular-weight immunoglobulin-binding protein from Yersinia pseudotuberculosis.

A low-molecular-weight immunoglobulin-binding protein (IBP) bound with the cell envelope has been isolated from Yersinia pseudotuberculosis cells and partially characterized. This IBP is a hydrophilic protein with a high polarity index of 55.3%. The molecular weight of the protein has been determined by MALDI-TOF mass spectrometry as 14.3 kD. CD spectroscopy showed that the IBP has high contents of the β-structure and random coil structure. The IBP contains glycine as the N-terminal amino acid. The protein can be stored for a long time at acidic pH values but aggregates and loses activity at alkaline and neutral pH. The IBP binds rabbit IgG with optimum at pH of 6.0–7.5. The IBP interacts with IgG molecule in the Fc-fragment region. The protein retains activity after heating at 100°C in the presence of SDS.

Formation of soluble chitosan–carrageenan polyelectrolyte complexes

The formation process of soluble chitosan (C)/κ-carrageenan (K) complexes was studied. It was shown that soluble complexes were obtained preferentially by mixing the starting components at given ratios whereas soluble complexes were formed only over a narrow range of starting-component ratios by titration of K with C. The formation of C/K polyelectrolyte complexes was confirmed by gel-permeation chromatography and centrifugation in a Percoll gradient. The formation process of C/K complexes depended on the C molecular weight and the concentrations and ratios of starting polysaccharides. It was shown that unbound components in addition to the complex remained in the mixture upon mixing carrageenan with high-molecular-weight C (C-HM) in 1:1.5 and 1:30 ratios (mol/mol) whereas low-molecular-weight C (C-LM) was bonded completely to the polyanion.

Biogenesis of β-barrel integral proteins of bacterial outer membrane

Gram-negative bacteria are enveloped by two membranes, the inner (cytoplasmic) (CM) and the outer (OM). The majority of integral outer membrane proteins are arranged in β-barrels of cylindrical shape composed of amphipathic antiparallel β-strands. In bacteria, β-barrel proteins function as water-filled pores, active transporters, enzymes, receptors, and structural proteins. Proteins of bacterial OM are synthesized in the cytoplasm as unfolded polypeptides with an N-terminal sequence that marks them for transport across the CM. Precursors of membrane proteins move through the aqueous medium of the cytosol and periplasm under the protection of chaperones (SecB, Skp, SurA, and DegP), then cross the CM via the Sec system composed of a polypeptide-conducting channel (SecYEG) and ATPase (SecA), the latter providing the energy for the translocation of the pre-protein. Pre-protein folding and incorporation in the OM require the participation of the Bam-complex, probably without the use of energy. This review summarizes current data on the biogenesis of the β-barrel proteins of bacterial OM. Data on the structure of the proteins included in the multicomponent system for delivery of the OM proteins to their destination in the cell and on their complexes with partners, including pre-proteins, are pre-sented. Molecular models constructed on the basis of structural, genetic, and biochemical studies that describe the mechanisms of β-barrel protein assembly by this molecular transport machinery are also considered.

Polysaccharide structure of tetrasporic red seaweed Tichocarpus crinitus

Sulfated polysaccharide isolated from tetrasporic plants of Tichocarpus crinitus was investigated. The polysaccharide was isolated by two methods: with water extraction at 80 °C (HT) and with a mild alkaline extraction (AE). The extracted polysaccharides were presented by non-gelling ones only, while galactose and 3,6-AG were the main monosaccharides, at the same time amount of 3,6-AG in AE polysaccharides was the similar to that of HT. According to methods of spectroscopy and mass spectrometry, the polysaccharide from tetrasporic T. crinitus contains main blocks of 1,3-linked β-D-galactopyranosyl-2,4-disulfates and 1,4-linked 3,6-anhydro-α-D-galactopyranosyl while 6-sulfated 4-linked galactopyranosyl resudies are randomly distributed along the polysaccharide chain. The alkaline treatment of HT polysaccharide results in obtaining polysaccharide with regular structure that composed of alternating 1,3-linked β-D-galactopyranosyl-2,4-disulfates and 1,4-linked 3,6-anhydro-α-D-galactopyranosyl residues. Native polysaccharide (HT) possessed both high anticoagulant and antiplatelet activity measured by fibrin clotting and platelet aggregation induced by collagen. This activity could be connected with peculiar chemical structure of HT polysaccharide which has high sulfation degree and contains also 3,6-anhydrogalactose in the polymer chain.