Yanhe Ma

Thermoanaerobacter tengcongensis sp. nov., a novel anaerobic, saccharolytic, thermophilic bacterium isolated from a hot spring in Tengcong, China.

A new, extremely thermophilic bacterium, designated strain MB4T, was isolated from a Chinese hot spring. The new isolate was an obligately anaerobic, rod-shaped, gram-negative, saccharolytic bacterium. Spore formation was not observed. Growth occurred at temperatures between 50 and 80 degrees C, with an optimum of around 75 degrees C; at pH values between 5.5 and 9.0, with an optimum of 7.0-7.5; and at salinities between 0 and 2.5% NaCl, with an optimum of around 0.2% NaCl. The organism utilized glucose, galactose, maltose, cellobiose, mannose, fructose, lactose, mannitol and starch. Acetate was the main end product from glucose fermentation. Thiosulfate and sulfur were reduced to hydrogen sulfide. Sulfate, sulfite and nitrate were not reduced. Growth was inhibited by hydrogen. The G+C content of the DNA was 33 mol%. Phylogenetic analyses based on the 16S rDNA sequence indicated that the isolate was a new member of the genus Thermoanaerobacter and formed a monophyletic unit within the Thermoanaerobacter cluster. Based on its phenotypic and phylogenetic characteristics, the isolate was proposed as a new species, Thermoanaerobacter tengcongensis. The type strain is MB4T (= Chinese Collection of Microorganisms AS 1.2430T = JCM 11007T).

Engineering central metabolic modules of Escherichia coli for improving β-carotene production.

ATP and NADPH are two important cofactors for production of terpenoids compounds. Here we have constructed and optimized β-carotene synthetic pathway in Escherichia coli, followed by engineering central metabolic modules to increase ATP and NADPH supplies for improving β-carotene production. The whole β-carotene synthetic pathway was divided into five modules. Engineering MEP module resulted in 3.5-fold increase of β-carotene yield, while engineering β-carotene synthesis module resulted in another 3.4-fold increase. The best β-carotene yield increased 21%, 17% and 39% after modulating single gene of ATP synthesis, pentose phosphate and TCA modules, respectively. Combined engineering of TCA and PPP modules had a synergistic effect on improving β-carotene yield, leading to 64% increase of β-carotene yield over a high producing parental strain. Fed-batch fermentation of the best strain CAR005 was performed, which produced 2.1g/L β-carotene with a yield of 60mg/g DCW.

Proteome Reference Map and Comparative Proteomic Analysis between a Wild Type Clostridium acetobutylicum DSM 1731 and its Mutant with Enhanced Butanol Tolerance and Butanol Yield

The solventogenic bacterium Clostridium acetobutylicum is an important species of the Clostridium community. To develop a fundamental tool that is useful for biological studies of C. acetobutylicum, we established a high resolution proteome reference map for this species. We identified 1206 spots representing 564 different proteins by mass spectrometry, covering approximately 50% of major metabolic pathways. To better understand the relationship between butanol tolerance and butanol yield, we performed a comparative proteomic analysis between the wild type strain DSM 1731 and the mutant Rh8, which has higher butanol tolerance and higher butanol yield. Comparative proteomic analysis of two strains at acidogenic and solventogenic phases revealed 102 differentially expressed proteins that are mainly involved in protein folding, solvent formation, amino acid metabolism, protein synthesis, nucleotide metabolism, transport, and others. Hierarchical clustering analysis revealed that over 70% of the 102 differentially expressed proteins in mutant Rh8 were either upregulated (e.g., chaperones and solvent formation related) or downregulated (e.g., amino acid metabolism and protein synthesis related) in both acidogenic and solventogenic phase, which, respectively, are only upregulated or downregulated in solventogenic phase in the wild type strain. This suggests that Rh8 cells have evolved a mechanism to prepare themselves for butanol challenge before butanol is produced, leading to an increased butanol yield. This is the first report on the comparative proteome analysis of a mutant strain and a base strain of C. acetobutylicum. The fundamental proteomic data and analyses will be useful for further elucidating the biological mechanism of butanol tolerance and/or enhanced butanol production.

Non-sterilized fermentative production of polymer-grade L-lactic acid by a newly isolated thermophilic strain Bacillus sp. 2-6.

Background The demand for lactic acid has been increasing considerably because of its use as a monomer for the synthesis of polylactic acid (PLA), which is a promising and environment-friendly alternative to plastics derived from petrochemicals. Optically pure l-lactic acid is essential for polymerization of PLA. The high fermentation cost of l-lactic acid is another limitation for PLA polymers to compete with conventional plastics. Methodology/Principal Findings A Bacillus sp. strain 2–6 for production of l-lactic acid was isolated at 55°C from soil samples. Its thermophilic characteristic made it a good lactic acid producer because optically pure l-lactic acid could be produced by this strain under open condition without sterilization. In 5-liter batch fermentation of Bacillus sp. 2–6, 118.0 g/liter of l-lactic acid with an optical purity of 99.4% was obtained from 121.3 g/liter of glucose. The yield was 97.3% and the average productivity was 4.37 g/liter/h. The maximum l-lactic acid concentration of 182.0 g/liter was obtained from 30-liter fed-batch fermentation with an average productivity of 3.03 g/liter/h and product optical purity of 99.4%. Conclusions/Significance With the newly isolated Bacillus sp. strain 2–6, high concentration of optically pure l-lactic acid could be produced efficiently in open fermentation without sterilization, which would lead to a new cost-effective method for polymer-grade l-lactic acid production from renewable resources.

Designing and creating a modularized synthetic pathway in cyanobacterium Synechocystis enables production of acetone from carbon dioxide

Ketones are a class of important organic compounds. As the simplest ketone, acetone is widely used as solvents or precursors for industrial chemicals. Presently, million tonnes of acetone is produced worldwide annually, from petrochemical processes. Here we report a biotechnological process that can produce acetone from CO(2), by designing and creating a modularized synthetic pathway in engineered cyanobacterium Synechocystis sp. PCC 6803. The engineered Synechocystis cells are able to produce acetone (36.0 mgl(-1) culture medium) using CO(2) as the sole carbon source, thus opens the gateway for biosynthesis of ketones from CO(2).

Efficient production of L-lactic acid from corncob molasses, a waste by-product in xylitol production, by a newly isolated xylose utilizing Bacillus sp. strain

Lignocellulosic biomass-derived sugars are considered nowadays to be an economically attractive carbohydrate feedstock for large-scale fermentations of bulk chemicals such as lactic acid. In the present study, corncob molasses containing a high content of xylose, which is one of the lignocellulosic biomasses and a waste by-product from xylitol production, was used for L-lactic acid production via a newly isolated xylose utilizing Bacillus sp. strain XZL9. Bacillus sp. strain XZL9 can utilize the mixture of sugars including xylose, arabinose, and glucose in corncob molasses for L-lactic acid production. High concentration of L-lactic acid (74.7 g l⁻¹) was obtained from corncob molasses (initial total sugars of 91.4 g l⁻¹) in fed-batch fermentation. This study provides an encouraging means of producing L-lactic acid from lignocellulosic resource such as the low-cost corncob molasses.

Microbial Biogeography of Six Salt Lakes in Inner Mongolia, China, and a Salt Lake in Argentina

ABSTRACT We used cultivation-independent methods to investigate the prokaryotic biogeography of the water column in six salt lakes in Inner Mongolia, China, and a salt lake in Argentina. These lakes had different salt compositions and pH values and were at variable geographic distances, on both local and intercontinental scales, which allowed us to explore the microbial community composition within the context of both contemporary environmental conditions and geographic distance. Fourteen 16S rRNA gene clone libraries were constructed, and over 200 16S rRNA gene sequences were obtained. These sequences were used to construct biotic similarity matrices, which were used in combination with environmental similarity matrices and a distance matrix in the Mantel test to discover which factors significantly influenced biotic similarity. We showed that archaeal biogeography was influenced by contemporary environmental factors alone (Na+, CO32−, and HCO3− ion concentrations; pH; and temperature). Bacterial biogeography was influenced both by contemporary environmental factors (Na+, Mg2+, and HCO3− ion concentrations and pH) and by geographic distance.

Medium optimization by combination of response surface methodology and desirability function: an application in glutamine production

An optimization strategy based on desirability function approach (DFA) together with response surface methodology (RSM) has been used to optimize production medium in L-glutamine fermentation. Fermentation problems often force to reach a compromise between different experimental variables in order to achieve the most suitable strategy applying in industrial production. The importance of the use of multi-objective optimization methods lies in the ability to cope with this kind of problems. A sequential RSM with different combinations of glucose and (NH4)2SO4 was performed to attain the optimal medium (OM-1) in glutamine production. Based on the result of RSM and the evaluation of production cost, a more economical optimal medium (OM-2) was obtained with the aid of DFA. In DFA study, glutamate, the main by-product in glutamine fermentation as another response was considered. Compared with OM-1 in validated experiment, similar amounts of glutamine were obtained in OM-2 while the concentration of glutamate and the production cost decreased by 53.6 and 7.1%, respectively.

Halophiles 2010: Life in Saline Environments

The world of halophilic microorganisms is highly diverse. Microbes adapted to life at high salt concentrations are found in all three domains of life: Archaea , Bacteria , and Eucarya . In some ecosystems salt-loving microorganisms live in such large numbers that their presence can be recognized

Activating transhydrogenase and NAD kinase in combination for improving isobutanol production.

Isobutanol is an excellent alternative biofuel. Fermentative production of isobutanol had been realized in several microorganisms by combining branched-chain amino acids synthetic pathway and Ehrlich pathway. In contrast to using plasmid overexpression and inducible promoters, genetically stable Escherichia coli strains for isobutanol production were constructed in this work by integrating essential genes into chromosome. A chromosome-based markerless gene modulation method was then developed for fine-tuning gene expression with multiple regulatory parts to improve isobutanol production. There was also a cofactor imbalance problem for anaerobic isobutanol synthesis. NADPH is the reducing equivalent required for isobutanol production, while the common reducing equivalent under anaerobic condition is NADH. Two strategies were used to modulate expression of transhydrogenase (pntAB) and NAD kinase (yfjB) genes to increase NADPH supply for improving isobutanol production. Plasmid overexpression of pntAB and yfjB genes either individually or in combination had little effect on isobutanol production. In contrast, modulating pntAB and yfjB gene expression in chromosome with multiple regulatory parts identified optimal modulators under aerobic and anaerobic conditions, respectively, and improved isobutanol production. Modulating pntAB gene alone led to 20% and 8% increase of anaerobic isobutanol titer and yield. Although modulating yfjB gene alone had nearly no effect, modulating pntAB and yfjB genes in combination led to 50% and 30% increase of isobutanol titer and yield in comparison with modulating pntAB gene alone. It was also found that increasing pntAB gene expression alone had a threshold for improving anaerobic isobutanol production, while activating NAD kinase could break through this threshold, leading to a yield of 0.92mol/mol. Our results suggested that transhydrogenase and NAD kinase had a synergistic effect on increasing NADPH supply and improving anaerobic isobutanol production. This strategy will be useful for improving production of target compounds using NADPH as reducing equivalent within their synthetic pathways. In addition, combined activation of PntAB and YfjB led to 28% and 22% increase of aerobic isobutanol titer and yield, resulting in production of 10.8g/L isobutanol in 24h with a yield of 0.62mol/mol.