Characterization of O-acetyl-(4-O-methylglucurono)xylan isolated from birch and beech

The structures of water-soluble birch and beech xylans, extracted from holocellulose using dimethyl sulfoxide, were determined employing 1H and 13C NMR spectroscopy together with chemical analysis. These polysaccharides were found to be O-acetyl-(4-O-methylglucurono)xylans containing one 4-O-methylglucuronic acid substituent for approximately every 15 D-xylose residues. The average degree of acetylation of the xylose residues in these polymers was 0.4. The presence of the structural element -->4)[4-O-Me-alpha-D-GlcpA-(1-->2)][3-O-Ac]-beta-D-Xylp-(1--> was demonstrated. Additional acetyl groups were present as substituents at C-2 and/or C-3 of the xylopyranosyl residues. Utilizing size-exclusion chromatography in combination with mass spectroscopy, the weight-average molar masses (and polydispersities) were shown to be 8000 (1.09) and 11,100 (1.08) for birch and beech xylan, respectively.     

Significant levels of extracellular reactive oxygen species produced by brown rot basidiomycetes on cellulose

It is often proposed that brown rot basidiomycetes use extracellular reactive oxygen species (ROS) to accomplish the initial depolymerization of cellulose in wood, but little evidence has been presented to show that the fungi produce these oxidants in physiologically relevant quantities. We used [(14)C]phenethyl polyacrylate as a radical trap to estimate extracellular ROS production by two brown rot fungi, Gloeophyllum trabeum and Postia placenta, that were degrading cellulose. Both fungi oxidized aromatic rings on the trap to give monohydroxylated and more polar products in significant yields. All of the cultures contained 2,5-dimethoxyhydroquinone, a fungal metabolite that has been shown to drive Fenton chemistry in vitro. These results show that extracellular ROS occur at significant levels in cellulose colonized by brown rot fungi, and suggest that hydroquinone-driven ROS production may contribute to decay by diverse brown rot species.     

Mutations of basic arginine residue 334 in the D1 protein of Photosystem II lead to unusual S2 state properties in Synechocystis sp. PCC 6803

The C-terminus region of the D1 protein of Photosystem II (PS II) is situated on the lumenal side of the complex and is likely to be involved in the coordination of the active site Mn atoms of the water oxidation complex (WOC). The strictly conserved arginine at position 334 (D1-334) was targeted for site-directed mutagenesis to explore the hypothesis that it is involved in the PS II extrinsic protein binding, chloride binding, or proton transfer. Although it was found that D1-R334 probably not essential for these functions, mutations at this position were found to uniquely alter the kinetics of S-state cycling in general and the properties of the S₂ state in particular. Substitutions of a glutamate (D1-R334E) and a valine (D1-R334V) for D1-R334 lead to an unusually stable (t _1/2 >30 min at room temp) S₂ state, but not S₃, as measured by double flash measurements on the bare platinum electrode. However, measurements of fluorescence decay in the presence of DCMU suggest the S₂ state is only modestly affected by the mutations. Possible reasons for these apparently contradictory results are discussed. This revised version was published online in June 2006 with corrections to the Cover Date.     

Enzyme-induced covalent modification of methionyl-tRNA synthetase from Bacillus stearothermophilus by methionyl-adenylate: identification of the labeled amino acid residues by matrix-assisted laser desorption-ionization mass spectrometry

Methionyl-tRNA synthetase (MetRS) from Bacillus stearothermophilus was shown to undergo covalent methionylation by a donor methionyl-adenylate, the mixed carboxylic-phosphoric acid anhydride synthesized by the enzyme itself. Covalent reaction of methionyl-adenylate with the synthetase or other proteins proceeds through the formation of an isopeptide bond between the carboxylate of the amino acid and the -NH₂ group of lysyl residues. The stoichiometries of labeling, as followed by TCA precipitation, were 2.2 ± 0.1 and 4.3 ± 0.1 mol of [¹⁴C]Met incorporated by 1 mol of the monomeric MS534 and the native dimeric species of B. stearo methionyl-tRNA synthetase, respectively. Matrix-assisted laser desorption-ionization mass spectrometry designated lysines-261, -295, -301 and -528 (or -534) of truncated methionyl-tRNA synthetase as the target residues for covalent binding of methionine. By analogy with the 3D structure of the monomeric M547 species of E. coli methionyl-tRNA synthetase, lysines-261, -295, and -301 would be located in the catalytic crevice of the thermostable enzyme where methionine activation and transfer take place. It is proposed that, once activated by ATP, most of the methionine molecules react with the closest reactive lysyl residues.     

Tryptophanyl fluorescence lifetime distribution of hyperthermophilic beta-glycosidase from molecular dynamics simulation: a comparison with the experimental data

A molecular dynamics simulation approach has been utilized to understand the unusual fluorescence emission decay observed for beta-glycosidase from the hyperthermophilic bacterium Solfolobus sulfotaricus (Sbeta gly), a tetrameric enzyme containing 17 tryptophanyl residues for each subunit. The tryptophanyl emission decay of Sbeta gly results from a bimodal distribution of fluorescence lifetimes with a short-lived component centered at 2.5 ns and a long-lived one at 7.4 ns (Bismuto E, Nucci R, Rossi M, Irace G, 1999, Proteins 27:71-79). From the examination of the trajectories of the side chains capable of causing intramolecular quenching for each tryptophan microenvironment and using a modified Stern-Volmer model for the emission quenching processes, we calculated the fluorescence lifetime for each tryptophanyl residue of Sbeta gly at two different temperatures, i.e., 300 and 365 K. The highest temperature was chosen because in this condition Sbeta gly evidences a maximum in its catalytic activity and is stable for a very long time. The calculated lifetime distributions overlap those experimentally determined. Moreover, the majority of trytptophanyl residues having longer lifetimes correspond to those originally identified by inspection of the crystallographic structure. The tryptophanyl lifetimes appear to be a complex function of several variables, such as microenvironment viscosity, solvent accessibility, the chemical structure of quencher side chains, and side-chain dynamics. The lifetime calculation by MD simulation can be used to validate a predicted structure by comparing the theoretical data with the experimental fluorescence decay results.     

Flatworm phylogeny from partial 18S rDNA sequences

Partial 18S rDNA sequences from 29 flatworms and 2 outgroup taxa were used in a cladistic analysis of the Platyhelminthes. Support for the clades in the resulting single most parsimonious tree was estimated through bootstrap analysis, jack-knife analysis and decay indices. The Acoelomorpha (Acoela and Nemertodermatida) were absent from the most parsimonious tree. The Acoela and the Fecampiidae form a strongly supported clade, the sister group of which may be the Tricladida. There is some support for monophyly of the rhabdocoel taxon Dalyellioida, previously regarded as paraphyletic. The sister group of the Neodermata remains unresolved. This revised version was published online in July 2006 with corrections to the Cover Date.     

Perspectives in the glycosciences – matrix-assisted laser desorption/ionization (MALDI) mass spectrometry of carbohydrates

The technique of matrix-assisted laser desorption/ionization (MALDI) mass spectrometry is reviewed with emphasis on the best type of matrix to use with carbohydrates, ways to obtain fragmentation and use of the technique for carbohydrate sequencing. © 1998 Rapid Science Ltd     

Acetic Acid, Ethanol and Steam Effects on the Growth of Botrytis cinerea in vitro and Combination of Steam and Modified Atmosphere Packaging to Control Decay in Kiwifruit

The effects of acetic acid fumigation, ethanol fumigation, and steam heat treatment on growth of Botrytis cinerea in vitro were investigated. The effect of steam heat treatments in combination with modified atmosphere packaging (MAP) on Botrytis decay development on 'Hayward' kiwifruit was also studied. The fungus was grown in Petri dishes on potato dextrose agar. Ethanol fumigation with 100  μ l/l for 3 or 6 min, or 200  μ l/l for 6 min enhanced the growth of B. cinerea . The effect of acetic acid on growth of B. cinerea was time and dosage-dependent. Fumigation with 1  μ l/l for 6 min, 2  μ l/l for 3 min, and 4  μ l/l for 3 min promoted radial growth of the fungus when compared to the growth of the untreated control. Fumigation with 2  μ l/l for 6 min delayed the growth of the fungus for the first 6 days, while fumigation with 6  μ l/l for 3 min delayed the growth of the fungus after the sixth day. Fumigation with 4 or 6  μ l/l acetic acid for 6 min, and 8  μ l/l acetic acid for 3 or 6 min resulted in complete inhibition of fungal growth. Steam heat treatment at 45°C for 6 min, and at 48, 51, and 54°C for 3 or 6 min completely inhibited fungal growth in vitro . Furthermore, steam treatments at 47, 50, and 53°C for 3 or 6 min completely inhibited decay at the stem end of kiwifruit kept at 10°C in MAP for 12 days. However, none of the steam treatments inhibited decay in wounds on the surface of the fruit kept in MAP.     

Long-term patterns of mass loss during the decomposition of leaf and fine root litter: an intersite comparison

Decomposition is a critical process in global carbon cycling. During decomposition, leaf and fine root litter may undergo a later, relatively slow phase; past long-term experiments indicate this phase occurs, but whether it is a general phenomenon has not been examined. Data from Long-term Intersite Decomposition Experiment Team, representing 27 sites and nine litter types (for a total of 234 cases) was used to test the frequency of this later, slow phase of decomposition. Litter mass remaining after up to 10 years of decomposition was fit to models that included (dual exponential and asymptotic) or excluded (single exponential) a slow phase. The resultant regression equations were evaluated for goodness of fit as well as biological realism. Regression analysis indicated that while the dual exponential and asymptotic models statistically and biologically fit more of the litter type–site combinations than the single exponential model, the latter was biologically reasonable for 27–65% of the cases depending on the test used. This implies that a slow phase is common, but not universal. Moreover, estimates of the decomposition rate of the slowly decomposing component averaged 0.139–0.221 year⁻¹ (depending on method), higher than generally observed for mineral soil organic matter, but one-third of the faster phase of litter decomposition. Thus, this material may be slower than the earlier phases of litter decomposition, but not as slow as mineral soil organic matter. Comparison of the long-term integrated decomposition rate (which included all phases of decomposition) to that for the first year of decomposition indicated the former was on average 75% that of the latter, consistent with the presence of a slow phase of decomposition. These results indicate that the global store of litter estimated using short-term decomposition rates would be underestimated by at least one-third.