Myocardium of type 2 diabetic and obese patients is characterized by alterations in sphingolipid metabolic enzymes but not by accumulation of ceramide

Data from animal experiments strongly suggest that ceramide is an important mediator of lipotoxicity in the heart and that accumulation of ceramide contributes to cardiomyocyte apoptosis associated with type 2 diabetes and obesity. However, it remains unknown whether a similar relationship is present also in the human heart. Therefore, we aimed to examine whether myocardial apoptosis in obese and type 2 diabetic patients is associated with elevated ceramide level. The study included 11 lean and 26 overweight or moderately obese subjects without (n = 11, OWT) or with (n = 15, T2D-OWT) a history of type 2 diabetes. Samples of the right atrial appendage were obtained from patients at the time of coronary bypass surgery. Compared with lean subjects, the extent of DNA fragmentation (a marker of apoptosis) was significantly higher in the myocardium of OWT patients and increased further in T2D-OWT subjects. However, the content of ceramide and sphingoid bases remained stable. Interestingly, the mRNA level of enzymes involved in synthesis and degradation of ceramide including serine palmitoyltransferase, sphingosine kinase 1, neutral sphingomyelinase, and ceramidases was markedly higher in the myocardium of OWT and T2D-OWT patients compared with lean subjects. Our results indicate that in the human heart, or at least in the atrium, ceramide is not a major factor in cardiomyocyte apoptosis associated with obesity and type 2 diabetes.     

Structural analysis of the lipid A isolated from Hafnia alvei 32 and PCM 1192 lipopolysaccharides

Hafnia alvei, a Gram-negative bacterium, is an opportunistic pathogen associated with mixed hospital infections, bacteremia, septicemia, and respiratory diseases. The majority of clinical symptoms of diseases caused by this bacterium have a lipopolysaccharide (LPS, endotoxin)-related origin. The lipid A structure affects the biological activity of endotoxins predominantly. Thus, the structure of H. alvei lipid A was analyzed for the first time. The major form, asymmetrically hexa-acylated lipid A built of β-d-GlcpN4P-(1→6)-α-d-GlcpN1P substituted with (R)-14:0(3-OH) at N-2 and O-3, 14:0(3-(R)-O-12:0) at N-2′, and 14:0(3-(R)-O-14:0) at O-3′, was identified by ESI-MSⁿ and MALDI-time-of-flight (TOF) MS. Comparative analysis performed by MS suggested that LPSs of H. alvei 32, PCM 1192, PCM 1206, and PCM 1207 share the identified structure of lipid A. LPSs of H. alvei are yet another example of enterobacterial endotoxins having the Escherichia coli-type structure of lipid A. The presence of hepta-acylated forms of H. alvei lipid A resulted from the addition of palmitate (16:0) substituting 14:0(3-OH) at N-2 of the α-GlcpN residue. All the studied strains of H. alvei have an ability to modify their lipid A structure by palmitoylation.     

Crystal structures of NodS N-methyltransferase from Bradyrhizobium japonicum in ligand-free form and as SAH complex

NodS is an S-adenosyl-l-methionine (SAM)-dependent N-methyltransferase that is involved in the biosynthesis of Nod factor (NF) in rhizobia, which are bacterial symbionts of legume plants. NF is a modified chitooligosaccharide (COS) signal molecule that is recognized by the legume host, where it initiates symbiotic processes leading to atmospheric nitrogen fixation. We report the crystal structure of recombinant NodS protein from Bradyrhizobium japonicum, which infects lupine and serradella legumes. Two crystal forms—ligand-free NodS and NodS in complex with S-adenosyl-l-homocysteine, which is a by-product of the methylation reaction—were obtained, and their structures were refined to resolutions of 2.43 Å and 1.85 Å, respectively. Although the overall fold (consisting of a seven-stranded β-sheet flanked by layers of helices) is similar to those of other SAM-dependent methyltransferases, NodS has specific features reflecting the unique character of its oligosaccharide substrate. In particular, the N-terminal helix and its connecting loop get ordered upon SAM binding, thereby closing the methyl donor cavity and shaping a long surface canyon that is clearly the binding site for the acceptor molecule. Comparison of the two structural forms of NodS suggests that there are also other conformational changes taking place upon the binding of the donor substrate. As an enzyme that methylates a COS substrate, NodS is the first example among all SAM-dependent methyltransferases to have its three-dimensional structure elucidated. Gaining insight about how NodS binds its donor and acceptor substrates helps to better understand the mechanism of NodS activity and the basis of its functional difference in various rhizobia.     

<Emphasis Type="Italic">Rhizobium leguminosarum</Emphasis> bv. <Emphasis Type="Italic">trifolii rosR</Emphasis> is required for interaction with clover,biofilm formation and adaptation to the environment

Background  

Rhizobium leguminosarum bv. trifolii is a symbiotic nitrogen-fixing bacterium that elicits nodules on roots of host plants Trifolium spp. Bacterial surface polysaccharides are crucial for establishment of a successful symbiosis with legumes that form indeterminate-type nodules, such as Trifolium, Pisum, Vicia, and Medicago spp. and aid the bacterium in withstanding osmotic and other environmental stresses. Recently, the R. leguminosarum bv. trifolii RosR regulatory protein which controls exopolysaccharide production has been identified and characterized.     

Complete lipopolysaccharide of Plesiomonas shigelloides O74:H5 (strain CNCTC 144/92). 2. Lipid A, its structural variability, the linkage to the core oligosaccharide, and the biological activity of the lipopolysaccharide

Plesiomonas shigelloides is a Gram-negative bacterium associated with waterborne infections, which is common in tropical and subtropical habitats. Contrary to the unified antigenic classification of P. shigelloides, data concerning the structure and activity of their lipopolysaccharides (LPS and endotoxin) are limited. This study completes the structural investigation of phenol- and water-soluble fractions of P. shigelloides O74 (strain CNCTC 144/92) LPS with the emphasis on lipid A heterogeneity, describing the entire molecule and some of its biological in vitro activities. Structures of the lipid A and the affinity-purified decasaccharide obtained by de-N,O-acylation of P. shigelloides O74 LPS were elucidated by chemical analysis combined with electrospray ionization multiple-stage mass spectrometry (ESI-MS(n)), MALDI-TOF MS, and NMR spectroscopy. Lipid A of P. shigelloides O74 is heterogeneous, and three major forms have been identified. They all were asymmetric, phosphorylated, and hexaacylated, showing different acylation patterns. The beta-GlcpN4P-(1-->6)-alpha-GlcpN1P disaccharide was substituted with the primary fatty acids: (R)-3-hydroxytetradecanoic acid [14:0(3-OH)] at N-2 and N-2' and (R)-3-hydroxydodecanoic acid [12:0(3-OH)] at O-3 and O-3'. The heterogeneity among the three forms (I-III) of P. shigelloides O74 lipid A was attributed to the substitution of the acyl residues at N-2' and O-3' with the secondary acyls: (I) cis-9-hexadecenoic acid (9c-16:1) at N-2' and 12:0 at O-3', (II) 14:0 at N-2' and 12:0 at O-3', and (III) 12:0 at N-2' and 12:0 at O-3'. The pro-inflammatory cytokine-inducing activities of P. shigelloides O74 LPS were similar to those of Escherichia coli O55 LPS.     

High frequency of drill holes in brachiopods from the Pliocene of Algeria and its ecological implications

The fossil record holds a wealth of ecological data, including data on biotic interactions. For example, holes in the skeletons of invertebrates produced by drilling activities of their enemies are widely used for exploring the intensity of such interactions through time because they are common and easily distinguished from non-biotic holes or holes produced by other types of interactions. Such drill holes have been described in numerous studies of Palaeozoic brachiopods but rarely in those focusing on brachiopods of the post-Palaeozoic, a striking pattern given that in the late Mesozoic and Cenozoic drilling gastropods diversified and frequencies of drilled molluscs increased dramatically. During the past several years, however, drilled brachiopods were reported in several studies of the Mesozoic and Cenozoic, suggesting that this phenomenon may be more common than has been previously assumed. Here we report on drilled brachiopods from a Pliocene locality in Algeria where 90 of 261 (34.5%) specimens of Megerlia truncata show evidence of predatory drilling. These data confirm that Cenozoic drilling frequencies of brachiopods may be locally high and, when taken together with other published data, that drilling frequencies are highly heterogeneous in space and time.     

Organogenesis of vegetative shoots from in vitro cultured flower buds of Mammillaria albicoma (Cactaceae)

We tested the organogenetic capacity of floral buds of Mammillaria albicoma Böed. (Cactaceae). Buds were incubated on solid MS medium supplemented with 0.1 mg l^?1 α-naphthaleneacetic acid (NAA) and 5.0 mg l^?1 6-benzylaminopurine (BA). Callus growth was observed from the cut explant base and from within the perianth. These calli during subsequent subcultures to the same medium gave rise to adventitious shoots. Shoots formed also directly from the perianth, as confirmed by observations in the light microscope and scanning electron microscope (SEM). On transfer to a fresh medium, the shoots produced proliferating cultures. This is the first report of regeneration of cactus shoots from floral explants.     

Chromium (III) Propionate and dietary fructans supplementation stimulate erythrocyte glucose uptake and beta-oxidation in lymphocytes of rats

The study describes the effects of 10-wk dietary supplementation with fructans (inulin and oligofructose, 5% and 10%, respectively) as well as the biomimetic Cr(III) propionate complex (0.5 and 5 mg Cr/kg diet) on blood glucose, insulin, glucose transmembrane transport, and β-oxidation of fatty acids in healthy male rats. No significant differences in blood serum glucose concentrations were found. Rats fed diets supplemented with the biomimetic complex (5 mg Cr/kg diet) had markedly decreased serum insulin level by 15%, whereas the red blood cells (RBCs) glucose transmembrane transport and β-oxidation of fatty acids in white blood cells (WBCs) were elevated by 9% and 77%, respectively. These effects were accompanied by a slight decrease of the insulin-resistance index. Oligofructose and the high-fructan diet (10%) were more effective in increasing the RBCs glucose transmembrane transport vs inulin and lowfructan diet (5%). Also, β-oxidation of fatty acids in WBCs was increased by 37.5% in groups fed the high-fructan diet (10%). The results suggest that dietary fructans and the biomimetic Cr(III) complex exerted beneficial effects on glucose and lipid metabolism, increasing the efficiency of their utilization.     

Conformational changes of L-rRNA during elongation of polypeptide

A ribosome undergoes significant conformational changes during elongation of a polypeptide chain, and these are correlated with structural changes of rRNAs. We tested 15 different oligonucleotides complementary to the selected, highly conserved seqences of rRNAs (L-rRNA, 5S rRNA and tRNA) important in protein biosynthesis. We carried out a reaction of binding Phe-tRNA to A site and a polymerization of polypeptide chains on the ribosomes converted either to pre- or to posttranslocational states. The inhibition of polymerization reaction by complementary oligonucleotides was high in all ribosomal states. The efficiency of inhibition of binding reaction was lower and more diverse than was the polypeptide elongation. We conclude that the selected oligonucleotides inhibit polypeptide synthesis with different effectivity, primarily depending on L-rRNA conformation within ribosome architecture.     

Enzymatic degradation studies of endomorphin-2 and its analogs containing N-methylated amino acids

In this paper, we describe the synthesis of novel endomorphin-2 analogs, containing N-methylated amino acids, consecutively in each position. The receptor-binding profile of the new analogs and their stability against enzymatic cleavage by commercially available peptidases, carboxypeptidase Y and aminopeptidase M, and a rat brain homogenate are reported. The best analog of this series, [Sar2]endomorphin-2, was almost equipotent with the parent peptide in the mu-receptor-binding assay and was also highly resistant to enzymatic degradation. This analog may be a suitable candidate for the in vivo antinociceptive studies.