Microbial degradation of octamethylcyclotetrasiloxane

The microbial degradation of low-molecular-weight polydimethylsiloxanes was investigated through laboratory experiments. Octamethylcyclotetrasiloxane was found to be biodegraded under anaerobic conditions in composted sewage sludge, as monitored by the occurrence of the main polydimethylsiloxane degradation product, dimethylsilanediol, compared to that found in experiments with sterilized control samples.     

Role of mycelium and extracellular protein in the biodegradation of 2,4,6-trichlorophenol by Phanerochaete chrysosporium.

The biodegradation of 2,4,6-trichlorophenol (2,4,6-TCP) by Phanerochaete chrysosporium was studied in batch systems. In experiments with mycelial suspension, the degradation of 2,4,6-TCP was found to occur in the absence of ligninase. Chloride ion was recovered in nearly stoichiometric amounts at the end of the process. The microorganism did not retain its degradation ability for more than 6 days under substrate-deficient conditions. Neither the mycelium nor the extracellular protein alone could degrade 2,4,6-TCP; both were required for complete degradation to occur. In experiments in which 2,4,6-TCP was exposed to the culture supernatant separated from its mycelium, negligible degradation was obtained and no chloride ion was recovered. No degradation was observed even when the supernatant was supplemented with hydrogen peroxide as a possible cosubstrate. In experiments performed with washed mycelium separated from its supernatant, no degradation took place until the mycelium released additional extracellular protein 5 to 6 h into the incubation. Additions of washed mycelium separated from its supernatant to active cultures also produced an increase in the rate of degradation in correspondence with the protein release. The protein release was independent of the presence of 2,4,6-TCP. The addition of cycloheximide to inhibit the synthesis of de novo proteins completely suppressed the release of protein by the mycelium and resulted in no 2,4,6-TCP degradation. Additions of culture supernatants containing a high concentration of extracellular protein to active cultures produced an increase in the rate of 2,4,6-TCP degradation.(ABSTRACT TRUNCATED AT 250 WORDS)     

Kinetics of biodegradation of p-nitrobenzoate and inhibition by benzoate in a pseudomonad.

The degradation of p-nitrobenzoate (p-NBA) by domestic sewage was inhibited by benzoate, and a model for this behavior was found in a soil isolate. The isolate, a pseudomonad, utilized p-NBA and benzoate by separate adaptive enzyme pathways. In oxygen uptake experiments, the degradation of p-NBA was competitively inhibited by benzoate, but the degradation of benzoate was not affected by the presence of p-NBA. 4-Nitrocatechol was not implicated in the inhibition. p-Hydroxybenzoate, which is the p-NBA degradation pathway, also had a decreased rate od degradation when benzoate was present. The growth rate of the isolate on the aromatic substrates and on glucose autoclaved in the medium was 0.3 h-1. When glucose was autoclaved separately, the growth rate was less, about 0.2 h-1. The apparent Km in oxygen uptake experiments was 25 micrometer for p-NBA and benzoate and 5 micrometer for p-hydroxybenzoate.     

Lab-Scale Biodegradation Study of BTEX under the Unsaturated Condition and Its Effect on Soil Matric Potential

Benzene, toluene, ethylbenzene, and xylene are collectively known as BTEX which contributes to volatile environmental contaminants. This present study investigates the microbial degradation of BTEX in batch and continuous soil column experiments and its effects on soil matric potential. Batch degradation experiments were performed with different initial concentrations of BTEX using the BTEX tolerant culture isolated from petroleum-contaminated soil. In batch study, the degradation pattern for single substrate showed that xylene was degraded much faster than other compounds followed by ethylbenzene, toluene, and benzene with the highest μ_max = 0.140 h^?1 during initial substrate concentration of 100 mg L^?1. Continuous degradation experiments were performed in a soil column with an inlet concentration of BTEX of about 2000 mg L^?1 under unsaturated flow in anaerobic condition. BTEX degradation pattern was studied with time and the matric potential of the soil at different parts along the length of the column were determined at the end of the experiment. In continuous degradation study, BTEX compounds were degraded with different degradation pattern and an increase in soil matric potential was observed with an increase in depth from top to bottom in the column with applied suction head. It was found that column biodegradation contributed to 69.5% of BTEX reduction and the bacterial growth increased the soil matric potential of about 34% on an average along the column height. Therefore, this study proves that it is significant to consider soil matric potential in modeling fate and transport of BTEX in unsaturated soils.     

Akt promotes increased mammalian cell size by stimulating protein synthesis and inhibiting protein degradation

Expression of constitutively active Akt3 was found to increase the size of MCF-7 cells approximately twofold both in vitro and in vivo. A regulatable version of Akt1 (MER-Akt) was also found capable of inducing a twofold increase in the size of H4IIE rat hepatoma cells. Rapamycin, a specific inhibitor of mTOR function, was found to inhibit the Akt-induced increase in cell size by 70%, presumably via inhibition of the Akt-induced increase in protein synthesis. To determine whether Akt could be inhibiting protein degradation, thereby contributing to its ability to induce an increase in cell size, we conducted protein degradation experiments in the H4IIE cell line. Activation of MER-Akt was found to inhibit protein degradation to a degree comparable to insulin treatment. The effects of these two agents on protein degradation were not additive, thereby suggesting that they were acting on a similar pathway. An inhibitor of the phosphatidylinositol 3-kinase pathway, LY-294002, blocked both insulin- and Akt-induced inhibition of protein degradation, again consistent with the hypothesis that both agents were acting on the same pathway. In contrast, rapamycin did not block the ability of either agent to inhibit protein degradation. These results indicate that Akt increases cell size through both mTOR-dependent and -independent pathways and that the latter involves inhibition of protein degradation. These studies are also consistent with the hypothesis that insulin's ability to regulate protein degradation is to a large extent mediated via Akt.     

Modeling dynamic experiments on the anaerobic degradation of molasses wastewater

The kinetics of anaerobic degradation of a molasses wastewater were measured under constant pH conditions in a laboratory scale packed bed reactor. In continuous and batch experiments the formation and degradation rates of the organic acids (butyric, propionic and acetic) have been followed. The influence of hydrogen gas on the acid degradation rates has been measured and, contrary to the literature and the thermo-dynamic calculations, no inhibition was detected, biofilm diffusional effects may be the reason. Two dynamic simulation models were tested, a heterogeneous model, which considered the biofilm diffusion-reaction phenomena and a quasihomogeneous model with the same kinetics. Except for hydrogen, the diffusion effects were found to be negligible. Otherwise both models gave essentially the same results and the time profiles of acids, hydrogen, carbon dioxide and methane agreed relatively well with dynamic startup experiments. Batch experiments showed the acid concentrations to be highly sensitive to the initial molasses concentration. This aspect was not included in the model but is being investigated further.     

Studies on the intracellular degradation of newly synthesized collagen in 3-methylcholanthrene induced fibrosarcoma cells.

The intracellular degradation of newly synthesized collagen was studied in both normal fibroblast and 3-methylcholanthrene induced fibrosarcoma cells. The degradation of newly synthesized collagen was examined using pulse-chase experiments and radioactive labelling techniques with [3H]-proline. The percentage of intracellular proteolysis of newly synthesized collagen was determined by measuring the formation of [3H]-hydroxyproline containing fragments in alcohol-soluble and insoluble fractions of normal cells and fibrosarcoma cells in the culture. The rate of degradation of newly formed collagen was then followed by estimating the radioactivity of [3H]-hydroxyproline at different intervals, during the chase period. The results clearly demonstrated that the percent of intracellular degradation of newly synthesized collagen was approximately three fold higher in fibrosarcoma cells than in normal fibroblast cells. The increased intracellular degradation of newly formed collagen was followed by an increase in the activity of cathepsin B and L in fibrosarcoma cells. The pulse-chase experiments indicated that the rate of degradation of newly synthesized collagen in fibrosarcoma cells is relatively greater than in normal fibroblast cells. In addition, as the labelling time increased, the formation of [3H]-hydroxyproline containing peptides in the ethanol-soluble fraction were found to be increased in both normal cells and fibrosarcoma cells, but the extent of formation was higher in fibrosarcoma cells compared to normal fibroblast cells. The results of this investigation collectively suggest that the intracellular degradation of newly synthesized collagen is enhanced in fibrosarcoma cells.     

Establishment and validation of computational model for MT1-MMP dependent ECM degradation and intervention strategies

MT1-MMP is a potent invasion-promoting membrane protease employed by aggressive cancer cells. MT1-MMP localizes preferentially at membrane protrusions called invadopodia where it plays a central role in degradation of the surrounding extracellular matrix (ECM). Previous reports suggested a role for a continuous supply of MT1-MMP in ECM degradation. However, the turnover rate of MT1-MMP and the extent to which the turnover contributes to the ECM degradation at invadopodia have not been clarified. To approach this problem, we first performed FRAP (Fluorescence Recovery after Photobleaching) experiments with fluorescence-tagged MT1-MMP focusing on a single invadopodium and found very rapid recovery in FRAP signals, approximated by double-exponential plots with time constants of 26 s and 259 s. The recovery depended primarily on vesicle transport, but negligibly on lateral diffusion. Next we constructed a computational model employing the observed kinetics of the FRAP experiments. The simulations successfully reproduced our FRAP experiments. Next we inhibited the vesicle transport both experimentally, and in simulation. Addition of drugs inhibiting vesicle transport blocked ECM degradation experimentally, and the simulation showed no appreciable ECM degradation under conditions inhibiting vesicle transport. In addition, the degree of the reduction in ECM degradation depended on the degree of the reduction in the MT1-MMP turnover. Thus, our experiments and simulations have established the role of the rapid turnover of MT1-MMP in ECM degradation at invadopodia. Furthermore, our simulations suggested synergetic contributions of proteolytic activity and the MT1-MMP turnover to ECM degradation because there was a nonlinear and marked reduction in ECM degradation if both factors were reduced simultaneously. Thus our computational model provides a new in silico tool to design and evaluate intervention strategies in cancer cell invasion.     

Importance of unattached bacteria and bacteria attached to sediment in determining potentials for degradation of xenobiotic organic contaminants in an aerobic aquifer.

The bacterial abundance, distribution, and degradation potential (in terms of degradation versus lack of degradation) for four xenobiotic compounds in an aerobic aquifer sediment have been examined in laboratory and field experiments. The xenobiotic compounds studied were benzene, toluene, o-xylene, and naphthalene (all at concentrations of approximately 120 micrograms/liter). The aerobic degradation experiments ran for approximately 90 days at 10 degrees C, which corresponded to the groundwater temperature. At the end of the experiment, the major part of the microbial biomass, quantified as acridine orange direct counts, was attached to the groundwater sediment (18 x 10(6) to 25 x 10(6) cells per g [dry weight], and only a minor part was unattached in the groundwater (0.6 x 10(6) to 5.5 x 10(6) cells per ml). Experiments involving aquifer sediment suspensions showed identical degradation potentials in the laboratory and in the field. However, laboratory experiments involving only groundwater (excluding aquifer sediment) showed less degradation potential than in situ experiments involving only groundwater, indicating that the manipulation or approach of the laboratory experiments could affect the determination of the degradation potentials. No differences were observed between the groundwater-only and the sediment compartments in the in situ experiments in the ability to degrade the compounds, but the maximum degradation rates were substantially lower in the groundwater-only compartment. Preparations used in laboratory experiments for studying the degradation potential for xenobiotic organic contaminants should contain sediment to obtain the highest numbers of bacteria as well as the broadest and most stable degradation.(ABSTRACT TRUNCATED AT 250 WORDS)     

Importance of unattached bacteria and bacteria attached to sediment in determining potentials for degradation of xenobiotic organic contaminants in an aerobic aquifer.

The bacterial abundance, distribution, and degradation potential (in terms of degradation versus lack of degradation) for four xenobiotic compounds in an aerobic aquifer sediment have been examined in laboratory and field experiments. The xenobiotic compounds studied were benzene, toluene, o-xylene, and naphthalene (all at concentrations of approximately 120 micrograms/liter). The aerobic degradation experiments ran for approximately 90 days at 10 degrees C, which corresponded to the groundwater temperature. At the end of the experiment, the major part of the microbial biomass, quantified as acridine orange direct counts, was attached to the groundwater sediment (18 x 10(6) to 25 x 10(6) cells per g [dry weight], and only a minor part was unattached in the groundwater (0.6 x 10(6) to 5.5 x 10(6) cells per ml). Experiments involving aquifer sediment suspensions showed identical degradation potentials in the laboratory and in the field. However, laboratory experiments involving only groundwater (excluding aquifer sediment) showed less degradation potential than in situ experiments involving only groundwater, indicating that the manipulation or approach of the laboratory experiments could affect the determination of the degradation potentials. No differences were observed between the groundwater-only and the sediment compartments in the in situ experiments in the ability to degrade the compounds, but the maximum degradation rates were substantially lower in the groundwater-only compartment. Preparations used in laboratory experiments for studying the degradation potential for xenobiotic organic contaminants should contain sediment to obtain the highest numbers of bacteria as well as the broadest and most stable degradation.(ABSTRACT TRUNCATED AT 250 WORDS)     

The influence of NaCl on the degradation rate of dichloromethane byHyphomicrobium sp.

The degradation of dichloromethane by the pure strainHyphomicrobium GJ21 and by an enrichment culture, isolated from a continuously operating biological trickling filter system, as well as the corresponding growth rates of these organisms were investigated in several batch experiments. By fitting the experimental data to generally accepted theoretical expressions for microbial growth, the maximum growth rates were determined. The effect of NaCl was investigated at salt concentrations varying from 0 to 1000 mM. Furthermore the dichloromethane degradation was investigated separately in experiments in which a high initial biomass concentration was applied. The results show that microbial growth is strongly inhibited by increased NaCl concentrations (50% reduction of _max at 200–250 mM NaCl), while a certain degree of adaptation has taken place within an operational system eliminating dichloromethane. A critical NaCl concentration for growth of 600 mM was found for the microbial culture isolated from an operational trickling filter, while a value of 375 mM was found for the pure cultureHyphomicrobium GJ21. The substrate degradation appears to be much less susceptible to inhibition by NaCl. Even at 800 mM NaCl relatively high substrate degradation rates are still observed, although this process is again dependent on the NaCl concentration. Here the substrate elimination is due to the maintenance requirements of the microorganisms. The inhibition of the dichloromethane elimination was also investigated in a laboratory scale trickling filter. The results of these experiments confirmed those obtained in the batch experiments. At NaCl concentrations exceeding 600 mM a considerable elimination of dichloromethane was still observed for during several months of operation. These observations indicate that the inhibition of microbial growth offers a significant control parameter against excessive biomass growth in biological trickling filters for waste gas treatment.     

Distinct energy requirement for nuclear import and export of importin beta in living cells

Carbohydrates with reactive aldehyde and ketone groups can undergo Maillard reactions with proteins to form advanced glycation end products. Oxalate monoalkylamide was identified as one of the advanced glycation end products formed from the Maillard reaction of ascorbate with proteins. In these experiments, we have analyzed human lens proteins immunochemically for the presence of oxalate monoalkylamide. Oxalate monoalkylamide was absent in most of the very young lenses but was present in old and cataractous lenses. The highest levels were found in senile brunescent lenses. Incubation experiments using bovine lens proteins revealed that oxalate monoalkylamide could form from the ascorbate degradation products, 2,3-diketogulonate and L-threose. These data provide the first evidence for oxalate monoalkylamide in vivo and suggest that ascorbate degradation and its binding to proteins are enhanced during lens aging and cataract formation.     

Structure of the O-polysaccharide of Providencia stuartii O4 containing 4-(N-acetyl-L-aspart-4-yl)amino-4,6-dideoxy-D-glucose

The O-polysaccharide of Providencia stuartii O4 was obtained by mild acid degradation of the lipopolysaccharide, and the following structure of the pentasaccharide repeating unit was established: [structure: see text] where D-Qui4N(L-AspAc) is 4-(N-acetyl-L-aspart-4-yl)amino-4,6-dideoxy-D-glucose, which has not been hitherto found in bacterial polysaccharides. Structural studies were performed using sugar and methylation analyses, Smith degradation and NMR spectroscopy, including conventional 2D 1H,1H COSY, TOCSY, NOESY and 1H,13C HSQC experiments as well as COSY and NOESY experiments run in an H(2)O-D(2)O mixture to reveal correlations for NH protons.     

Ribosome stacking defines CGS1 mRNA degradation sites during nascent peptide-mediated translation arrest

Expression of the Arabidopsis CGS1 gene that codes for cystathionine gamma-synthase is feedback-regulated at the step of mRNA degradation in response to S-adenosyl-L-methionine (AdoMet). This regulation occurs during translation and involves AdoMet-induced temporal translation arrest prior to the mRNA degradation. Here, we have identified multiple intermediates of CGS1 mRNA degradation with different 5' ends that are separated by approximately 30 nucleotides. Longer intermediates were found to be produced as the number of ribosomes loaded on mRNA was increased. Sucrose density gradient centrifugation experiments showed that the shortest mRNA degradation intermediate was associated with monosomes, whereas longer degradation intermediates were associated with multiple ribosomes. Immunoblot analyses revealed a ladder of premature polypeptides whose molecular weights corresponded to products of ribosomes in a stalled stack. An increase in smaller premature polypeptides was observed as the number of ribosomes loaded on mRNA increased. These results show that AdoMet induces the stacking of ribosomes on CGS1 mRNA and that multiple mRNA degradation sites probably correspond to each stacked ribosome.     

Structure of the O-specific polysaccharide of the lipopolysaccharide of Rahnella aquatilis 95 U003

The O-polysaccharide of Rahnella aquatilis 95 U003 was obtained by mild acid degradation of the lipopolysaccharide and studied by sugar and methylation analyses, Smith degradation and (1)H and (13)C NMR spectroscopy, including 2D (1)H,(1)H COSY, TOCSY, ROESY, H-detected (1)H,(13)C HSQC and HMQC-TOCSY experiments. The O-polysaccharide was found to have a branched hexasaccharide repeating unit of the following structure:     

Detection of very low saturation constants in anaerobic digestion: Influences of calcium carbonate precipitation and pH

Samples taken from a fluidized-bed reactor revealed very low saturation constants for the degradation of acetate (2–12 mg/l) and propionate (<3 mg/l). The higher values for the acetate degradation appear to be caused by mass-transport limitation due to calcium carbonate precipitation within the biofilm. The intrinsic saturation constant is about 3 mg/l, which is significantly lower than previously published values for pure and mixed cultures. The influence of the pH on the saturation constant was investigated in fed-batch experiments. Contrary to the hypothesis that only the undissociated acid is the effective substrate, no significant influence of pH on the saturation constant (given as concentration of total acid) was observed. Batch experiments with n-butyrate revealed hyperbolic progress curves, which might be misinterpreted as a sign of a high saturation constant. However, fed-batch experiments showed that, for n-butyrate degradation, the saturation constant is very low. The isomerisation to isobutyrate and other side-reactions, for which indications were found, influence the progress curve such that an elevated saturation constant will result as an artifact. Thus saturation constants for n-butyrate degradation obtained from batch experiments have to be viewed critically. Received: 17 October 1997 / Received revision: 19 January 1998 / Accepted: 24 January 1998     

Potential of preliminary test methods to predict biodegradation performance of petroleum hydrocarbons in soil

Preliminary tests at different scales such as degradation experiments (laboratory) in shaking flasks, soil columns and lysimeters as well as in situ respiration tests (field) were performed with soil from two hydrocarbon contaminated sites. Tests have been evaluated in terms of their potential to provide information on feasibility, degradation rates and residual concentration of bioremediation in the vadose zone. Sample size, costs and duration increased with experimental scale in the order shaking flasks – soil columns – lysimeter – in situ respiration tests, only time demand of respiration tests was relatively low. First-order rate constants observed in degradation experiments exhibited significant differences between both, different experimental sizes and different soils. Rates were in line with type and history of contamination at the sites, but somewhat overestimated field rates particularly in small scale experiments. All laboratory experiments allowed an estimation of residual concentrations after remediation. In situ respiration tests were found to be an appropriate pre-testing and monitoring tool for bioventing although residual concentrations cannot be predicted from in situ respiration tests. Moreover, this method does not account for potential limitations that might hamper biodegradation in the longer term but only reflects the actual degradation potential when the test is performed.     

嗜麦芽寡养单胞菌全基因组测序及降解荧蒽特性研究

随着当前工业化社会的发展,由多环芳烃等石油污染物造成的土壤污染已经成为了世界性的环境问题。本实验在天津滨海石油污染场地分离筛选出一株多环芳烃高效降解菌株W18,经过细胞形态、理化实验和分子生物学鉴定该菌株为嗜麦芽寡养单胞菌(Stenotrophomonas maltophilia)。在表面活性剂吐温80诱导下,该菌对荧蒽降解率可达到73%。利用细菌全基因组De novo测序技术发现,其包括1个环状拓扑结构,基因大小为4 738 432 bp,GC含量为66.69%。通过与主要数据库(COG,GO,KEGG)进行比对注释,发现W18菌株中有25个基因编码了与PAHs降解有关的加氧酶。嗜麦芽寡养单胞菌为土壤石油污染处理的研究提供菌种资源,对其降解功能基因的研究将为微生物-多环芳烃高效修复体系提供重要的理论依据。     

Anaerobic degradation of protocatechuate (3,4-dihydroxybenzoate) by Thauera aromatica strain AR-1

The denitrifying bacterium Thauera aromatica strain AR-1 grows anaerobically with protocatechuate (3,4-dihydroxybenzoate (DHB)) as sole energy and carbon source. This bacterium harbors two distinct pathways for degradation of aromatic compounds, the benzoyl-coenzyme A (CoA) pathway for benzoate degradation and the hydroxyhydroquinone (HHQ) pathway for degradation of 3,5-DHB. In order to elucidate whether protocatechuate is degraded via the benzoyl-CoA or the HHQ pathway, induction experiments were carried out. Dense suspensions of cells grown on protocatechuate or benzoate readily degraded benzoate and protocatechuate but not 3,5-DHB. Dense suspensions of 3,5-DHB-grown cells degraded 3,4- and 3,5-DHB at similar rates, but benzoate was not degraded. 3,5-DHB hydroxylating activity was found only in cells grown with this substrate. HHQ dehydrogenase activity was found in extracts of cells grown with 3,5-DHB and at a low rate also in protocatechuate-grown cells, but not in extracts of cells grown with benzoate. Activities of protocatechuyl-CoA synthetase and protocatechuyl-CoA reductase leading to 3-hydroxybenzoyl-CoA were found in extracts of cells grown with protocatechuate. There was no repression of the HHQ pathway by the presence of protocatechuate, unlike by degradation of benzoate. We conclude that protocatechuate is not degraded via the HHQ pathway because there was no evidence of a hydroxylation reaction involved in this process. Instead, our results strongly suggest that protocatechuate is degraded via a pathway which connects to the benzoyl-CoA route of degradation.     

Studies on the Amino Acids Fermentation of Pentose Materials

This work was undertaken to determine whether the degradation of sugars by Brevibacterium pentoso-aminoacidicum nov. sp., a bacterium capable of producing amino acids from pentoses and hexoses, was due to constitutive or inducible enzymes. It was also intended to clarify the reason for the substrate specificity in the fermentation of sugars by this bacterium. After a series of experiments using washed resting cells grown on various kinds of sugars or their cell-free extracts, it was found that the enzymes involved in the degradation of pentoses were inducible, while those of hexose metabolism were constitutive. The activities of several enzymes related to the pathways of pentose metabolism were demonstrated and the substrate specificity of sugar degradation by this strain was explained satisfactorily by the inducer specificity of these enzymes.