Molecular architecture of protein-RNA recognition sites

The molecular architecture of protein-RNA interfaces are analyzed using a non-redundant dataset of 152 protein-RNA complexes. We find that an average protein-RNA interface is smaller than an average protein-DNA interface but larger than an average protein–protein interface. Among the different classes of protein-RNA complexes, interfaces with tRNA are the largest, while the interfaces with the single-stranded RNA are the smallest. Significantly, RNA contributes more to the interface area than its partner protein. Moreover, unlike protein–protein interfaces where the side chain contributes less to the interface area compared to the main chain, the main chain and side chain contributions flipped in protein-RNA interfaces. We find that the protein surface in contact with the RNA in protein-RNA complexes is better packed than that in contact with the DNA in protein-DNA complexes, but loosely packed than that in contact with the protein in protein–protein complexes. Shape complementarity and electrostatic potential are the two major factors that determine the specificity of the protein-RNA interaction. We find that the H-bond density at the protein-RNA interfaces is similar with that of protein-DNA interfaces but higher than the protein–protein interfaces. Unlike protein-DNA interfaces where the deoxyribose has little role in intermolecular H-bonds, due to the presence of an oxygen atom at the 2′ position, the ribose in RNA plays significant role in protein-RNA H-bonds. We find that besides H-bonds, salt bridges and stacking interactions also play significant role in stabilizing protein-nucleic acids interfaces; however, their contribution at the protein–protein interfaces is insignificant.     

Simultaneous biological removal of nitrogen–sulfur–carbon: Recent advances and challenges

Biological removal of carbon, nitrogen and sulfur is drawing increasing research interest in search for an efficient and cost-effective wastewater treatment. While extensive work on separate removal of nitrogen and sulfur is well documented, investigation on simultaneous denitrifying sulfide removal has only been reported recently. Most of the work on denitrifying sulfide removal has been focusing on bioreactor performance, loading and operating conditions. Nonetheless, underlying principles elucidating the biochemical reactions and the mechanisms of the microbial degradation are yet to be established. In addition, unstable denitrifying sulfide removal which is a major operating problem that hinders practical application of the process, is yet to be resolved. This paper provides a review on the state-of-the-art development of simultaneous biological removal of sulfur, nitrogen and carbon. Research on bioreactor operation and performance, reactor configurations, mechanisms and modeling work including the use of mass balance analysis and artificial neural networks is delineated. An in-depth discussion on the microbial community and functional consortium is also provided. Challenges and future work on simultaneous biological removal of nitrogen–sulfur–carbon are also outlined.     

Removal of Contaminating Metals from Soil by Sulfur-Based Bioleaching and Biogenic Sulfide-Based Precipitation

The potential of a hybrid process incorporating sulfur-based bioleaching and sulfide-based precipitation for treatment of metal-contaminated soil was examined in batch-type experiments. The sulfur-based soil bioleaching process with Acidithiobacillus sp. could be initiated at a wide range of initial pH from 4.0 to 6.3. After 15 days, 98% of Zn, 89% of Cu and 79% of Cd was bioleached. The gaseous sulfides recycling from Desulfovibrio sp.-mediated sulfate-reducing reactor via N₂ sparging efficiently treated metal-loaded soil leachate. With a sulfide/metal ratio of 3.0, 88% of Zn, 100% of Cu and 95% of Cd were precipitated, resulting in effluent metal concentrations of 3.5 mg Zn²⁺/L, 0.2 mg Cu²⁺/L and 0.03 mg Cd²⁺/L.

Supplemental materials are available for this article. Go to the publisher's online edition of Geomicrobiology Journal to view the supplemental file.     

Formation of Covellite (CuS) Under Biological Sulfate-Reducing Conditions

Sulfate-reducing bacteria (SRB) play a major role in the precipitation of metal sulfides in the environment. In this work, biogenic copper sulfide formation was examined in cultures of SRB and compared to chemically initiated Cu sulfide precipitation as a reference system. Mixed cultures of SRB were incubated at 22, 45, and 60°C in nutrient solutions that contained copper sulfate. Abiotic reference samples were produced by reacting uninoculated liquid media with Na₂S solutions under otherwise identical conditions. Precipitates were collected anaerobically by centrifugation, frozen in liquid N₂, and freeze-dried, followed by analysis using X-ray diffraction (XRD), X-ray fluorescence, and scanning electron microscopy. Covellite (CuS) was the only mineral found in the precipitates. Covellite was less crystalline in the biogenic precipitates than in the abiotic samples based on XRD peak widths and peak to background ratios. Poor crystallinity may be the result of slower precipitation rates in bacterial cultures as compared to the abiotic reference systems. Furthermore, bacterial cells may inhibit the nucleation steps that lead to crystal formation. Incubation at elevated temperatures improved the crystallinity of the biotic specimens.     

Continuous hydrogen peroxide production by organic buffers in phytoplankton culture media

We investigated the production of hydrogen peroxide (HOOH) in illuminated seawater media containing a variety of zwitterionic buffers. Production rates varied extensively among buffers, with 4‐(2‐hydroxyethyl)1‐piperazineethanesulfonic acid (HEPES) highest and N‐Tris(hydroxymethyl)methyl‐3‐aminopropanesulfonic acid (TAPS) among the lowest. The rate of HOOH accumulation was remarkably consistent over many days, and increased linearly with buffer concentration, natural seawater concentration, and light level. Concentrations of HEPES commonly used in culture media (1–10 mM) generated enough HOOH to kill the axenic Prochlorococcus strain VOL1 during growth in enriched seawater media at lower, environmentally realistic cell concentrations and/or under high light exposure. We also demonstrated that HEPES can be used experimentally to study the biological effects of chronic exposure to sublethal levels of HOOH such as may be experienced by light‐exposed microorganisms.     

Antioxidative responses in Vitis vinifera infected by grapevine fanleaf virus

The antioxidative response of grapevine leaves (Vitis vinifera cv. Trebbiano) affected by the presence of grapevine fanleaf virus was studied during the summer of 2010 at three different harvest times (July 1st and 26th, and August 30th). At the first and second harvest, infected leaves showed increases in the concentration of superoxide radical and hydrogen peroxide, the latter increasing for enhanced activity of superoxide dismutase. In contrast, at the last harvest time, increases in the ascorbate pool and ascorbate peroxidase activity maintained hydrogen peroxide to control levels. The glutathione pool was negatively affected as summer progressed, showing a decrease in its total and reduced form amounts. At the same time, increases in the ascorbate pool were observed, making antioxidant defenses of grapevine effective also at the last harvest time. Increases in phenolic acids, and in particular in p-hydroxybenzoic acid, at the first and second harvest might have enhanced the efficiency of the antioxidant system through an interrelation between a peroxidase/phenol/ascorbate system and the NADPH/glutathione/ascorbate cycle. The lack of increase in p-hydroxybenzoic acid at the third harvest could be due instead to the enhanced utilization of this acid for hydrogen peroxide detoxification. With time, grapevine plants lost their capacity to contrast the spread of grapevine fanleaf virus, but acquired a greater ability to counteract pathogen-induced oxidative stress, being endowed with more reduced antioxidant pools.     

Phytuberol,from Japanese Domestic Tobacco,Nicotiana tabacum cv. Suifu

Carcinogenesis is believed to be induced through the oxidative damage of DNA, and antioxidants are expected to suppress it. So, the polyphenolic antioxidants in daily foods were investigated to see whether they protect against genetic damage by active oxygen. In the evaluation, we used a bioassay and a chemical determination, a Salmonella mutagenicity test for mutation by a N-hydroxyl radical from one of the dietary carcinogens 3-amino-1-methyl-5H-pyrido[4,3-b]indole and the formation of 8-hydroxyl (8-OHdG) from 2′-deoxyguanosine (2′-dG) in a Fenton OH-radical generating system. Thirty-one antioxidants including flavonoids were compared in terms of radical-trapping activity with bacterial DNA and 2′-dG. Antioxidants inhibited the mutation but the IC₅₀ values were in the mM order. Against 8-OHdG formation, only α-tocopherol had a suppressive effect with an IC₅₀ of 1.5 μM. Thus, except α-tocopherol, the dietary antioxidants did not scavenge the biological radicals faster than bacterial DNA and intact 2′-dG, indicating that they failed to prevent oxidative gene damage and probably carcinogenesis.     

Involvement of Lysosomal Cysteine Proteases in Hydrogen Peroxide-induced Apoptosis in HL-60 Cells

Hydrogen peroxide is a well-known mediator of apoptosis. As a mechanism for H₂O₂-induced apoptosis, both a mitochondrial Cyt.c-dependent pathway and a lysosome-mediated pathway have been suggested. However, the relative roles of and the relation between these two pathways in H₂O₂-induced apoptosis remain to be discovered. In this study, to find the relative roles of the lysosomal and mitochondrial pathways, the effects of E-64-d, a cell-permeable inhibitor of lysosomal cysteine proteases, on apoptosis caused by H₂O₂ in HL-60 cells were investigated. It was found that the concentration of H₂O₂ strongly affected the inhibitory effect of E-64-d on the apoptosis in HL-60 cells: dose-dependent inhibition (up to 40%) of both DNA fragmentation and caspase-3 activation was observed when a high concentration of H₂O₂ (50 μM) was used to induce apoptosis, but no inhibitory effect was detected when a low concentration (10 μM) was used. Consistent with these observations, apparent lysosomal destabilization was observed only with 50 μM H₂O₂. The release of mitochondrial Cyt.c, in contrast, was observed at both 10 μM and 50 μM. These results indicated that the mitochondrial Cyt.c-mediated pathway predominates in the H₂O₂-induced apoptosis in HL-60 cells and the lysosomal mediated pathway is partially involved when high concentrations of H₂O₂ are used to induce apoptosis.     

Stabilization of Flavobacterium meningosepticum Glycerol Kinase by Introduction of a Hydrogen Bond

The thermostability of Flavobacterium meningosepticum glycerol kinase was increased by the change from Ser329 to Asp [Protein Eng., 14, 663-667 (2001)]. Based on a three-dimensional structure model of the mutant, we have postulated that a new charged-neutral hydrogen bond was formed between Asp329 and Ser414, and the formation of the hydrogen bond contributed to the stabilization of the tertiary structure and increased thermostability of the mutant enzyme. If the postulation is the case, FGK thermostabilization would be possible similarly by the single amino acid substitution from Ser414 to another amino acid which could form the hydrogen bond with Ser329. We did a single amino acid substitution of the wild-type enzyme from Ser414 to Asn. As we expected, S414N showed comparable thermostability to that of S329D. On the other hand, a difference in kinetic properties for ATP between S414N and S329D was observed.     

The Heterogeneity of the 7S Soybean Protein by Sepharose Gel Chromatography and Disc Gel Electrophoresis

Two kinds of N-acetylmuramidase, M-1 and M-2 enzymes, that were isolated from the cultural broth of Stm. globisporus 1829, were remarkably different in amino acid composition, immunological properties and modes of lytic action from each other. The M-1 enzyme was composed of 186 amino acid residues of which two moles were of half cystine, while the M-2 enzyme was composed of 99 amino acid residues with no cysteine. The hydrolyzing action of the M-2 enzyme was suppressed by the presence of an O-acetyl group on muramic acid residues in the peptidoglycan moiety, while that of the M-l enzyme was independent of the presence of O-acetyl groups. However, the hydrolyzing activity of both enzymes was enhanced when some muramic acid residues were substituted with stem peptides containing alanine, isoglutamine and lysine.