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1.
Unlocking Nature's Biosynthetic Power-Metabolic Engineering for the Fermentative Production of Chemicals.
Hoff, B, Plassmeier, J, Blankschien, M, Letzel, AC, Kourtz, L, Schröder, H, Koch, W, Zelder, O
Angewandte Chemie (International ed. in English). 2021;(5):2258-2278
Abstract
Fermentation as a production method for chemicals is especially attractive, as it is based on cheap renewable raw materials and often exhibits advantages in terms of costs and sustainability. The tremendous development of technology in bioscience has resulted in an exponentially increasing knowledge about biological systems and has become the main driver for innovations in the field of metabolic engineering. Progress in recombinant DNA technology, genomics, and computational methods open new, cheaper, and faster ways to metabolically engineer microorganisms. Existing biosynthetic pathways for natural products, such as vitamins, organic acids, amino acids, or secondary metabolites, can be discovered and optimized efficiently, thereby enabling competitive commercial production processes. Novel biosynthetic routes can now be designed by the rearrangement of nature's unlimited number of enzymes and metabolic pathways in microbial strains. This expands the range of chemicals accessible by biotechnology and has yielded the first commercial products, while new fermentation technologies targeting novel active ingredients, commodity chemicals, and CO2 -fixation methods are on the horizon.
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2.
Less Unfavorable Salt Bridges on the Enzyme Surface Result in More Organic Cosolvent Resistance.
Cui, H, Eltoukhy, L, Zhang, L, Markel, U, Jaeger, KE, Davari, MD, Schwaneberg, U
Angewandte Chemie (International ed. in English). 2021;(20):11448-11456
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Abstract
Biocatalysis for the synthesis of fine chemicals is highly attractive but usually requires organic (co-)solvents (OSs). However, native enzymes often have low activity and resistance in OSs and at elevated temperatures. Herein, we report a smart salt bridge design strategy for simultaneously improving OS resistance and thermostability of the model enzyme, Bacillus subtilits Lipase A (BSLA). We combined comprehensive experimental studies of 3450 BSLA variants and molecular dynamics simulations of 36 systems. Iterative recombination of four beneficial substitutions yielded superior resistant variants with up to 7.6-fold (D64K/D144K) improved resistance toward three OSs while exhibiting significant thermostability (thermal resistance up to 137-fold, and half-life up to 3.3-fold). Molecular dynamics simulations revealed that locally refined flexibility and strengthened hydration jointly govern the highly increased resistance in OSs and at 50-100 °C. The salt bridge redesign provides protein engineers with a powerful and likely general approach to design OSs- and/or thermal-resistant lipases and other α/β-hydrolases.
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Shining New Light on the Kinetics of Water Uptake by Organic Aerosol Particles.
Diveky, ME, Gleichweit, MJ, Roy, S, Signorell, R
The journal of physical chemistry. A. 2021;(17):3528-3548
Abstract
The uptake of water vapor by various organic aerosols is important in a number of applications ranging from medical delivery of pharmaceutical aerosols to cloud formation in the atmosphere. The coefficient that describes the probability that the impinging gas-phase molecule sticks to the surface of interest is called the mass accommodation coefficient, αM. Despite the importance of this coefficient for the description of water uptake kinetics, accurate values are still lacking for many systems. In this Feature Article, we present various experimental techniques that have been evoked in the literature to study the interfacial transport of water and discuss the corresponding strengths and limitations. This includes our recently developed technique called photothermal single-particle spectroscopy (PSPS). The PSPS technique allows for a retrieval of αM values from three independent, yet simultaneous measurements operating close to equilibrium, providing a robust assessment of interfacial mass transport. We review the currently available data for αM for water on various organics and discuss the few studies that address the temperature and relative humidity dependence of αM for water on organics. The knowledge of the latter, for example, is crucial to assess the water uptake kinetics of organic aerosols in the Earth's atmosphere. Finally, we argue that PSPS might also be a viable method to better restrict the αM value for water on liquid water.
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Transcriptome analyses unravel CYP1A1 and CYP1B1 as novel biomarkers for disinfection by-products (DBPs) derived from chlorinated algal organic matter.
Wu, B, Hong, H, Xia, Z, Liu, H, Chen, X, Chen, J, Yan, B, Liang, Y
Journal of hazardous materials. 2020;:121685
Abstract
Disinfection by-products (DBPs) are generated during chlorination of drinking water. Previous studies demonstrate that DBPs are cytotoxic, genotoxic and associated with an increased risk of human cancer. However, the molecular basis of DBPs-induced toxic effects remains unclear. Here, we chlorinated samples of algal-derived organic matter (AOM) and sediment organic matter (SOM) from a local drinking water reservoir. Chemical properties, toxicities and transcriptomic profiles of human Caco-2 cell exposed to AOM and SOM were compared before and after chlorination. We analyzed chlorination-caused distinct gene expression patterns between AOM and SOM, and identified a set of 22 differentially expressed genes under chlorination of AOM that are different from chlorinated SOM. Consequent network analysis indicates that differential CYP1A1, CYP1B1, ID1 and ID2 are common targets of the upstream regulators predicted in the AOM group, but not the SOM group. Through experimental validation and data integration from previous reports related to DBPs or environmental stressors, we found that CYP1A1 and CYP1B1 are specifically up-regulated after chlorinating AOM. Our study demonstrates that the two CYP1 genes likely act as novel biomarkers of AOM derived DBPs, and this would be helpful for testing drinking water DBPs toxicity and further monitoring drinking water safety.
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Nutrimetabolomics reveals food-specific compounds in urine of adults consuming a DASH-style diet.
Reisdorph, NA, Hendricks, AE, Tang, M, Doenges, KA, Reisdorph, RM, Tooker, BC, Quinn, K, Borengasser, SJ, Nkrumah-Elie, Y, Frank, DN, et al
Scientific reports. 2020;(1):1157
Abstract
Although health benefits of the Dietary Approaches to Stop Hypertension (DASH) diet are established, it is not understood which food compounds result in these benefits. We used metabolomics to identify unique compounds from individual foods of a DASH-style diet and determined if these Food-Specific Compounds (FSC) are detectable in urine from participants in a DASH-style dietary study. We also examined relationships between urinary compounds and blood pressure (BP). Nineteen subjects were randomized into 6-week controlled DASH-style diet interventions. Mass spectrometry-based metabolomics was performed on 24-hour urine samples collected before and after each intervention and on 12 representative DASH-style foods. Between 66-969 compounds were catalogued as FSC; for example, 4-hydroxydiphenylamine was found to be unique to apple. Overall, 13-190 of these FSC were detected in urine, demonstrating that these unmetabolized food compounds can be discovered in urine using metabolomics. Although linear mixed effects models showed no FSC from the 12 profiled foods were significantly associated with BP, other endogenous and food-related compounds were associated with BP (N = 16) and changes in BP over time (N = 6). Overall, this proof of principle study demonstrates that metabolomics can be used to catalog FSC, which can be detected in participant urine following a dietary intervention.
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Experimental metatranscriptomics reveals the costs and benefits of dissolved organic matter photo-alteration for freshwater microbes.
Nalven, SG, Ward, CP, Payet, JP, Cory, RM, Kling, GW, Sharpton, TJ, Sullivan, CM, Crump, BC
Environmental microbiology. 2020;(8):3505-3521
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Abstract
Microbes and sunlight convert terrigenous dissolved organic matter (DOM) in surface waters to greenhouse gases. Prior studies show contrasting results about how biological and photochemical processes interact to contribute to the degradation of DOM. In this study, DOM leached from the organic layer of tundra soil was exposed to natural sunlight or kept in the dark, incubated in the dark with the natural microbial community, and analysed for gene expression and DOM chemical composition. Microbial gene expression (metatranscriptomics) in light and dark treatments diverged substantially after 4 h. Gene expression suggested that sunlight exposure of DOM initially stimulated microbial growth by (i) replacing the function of enzymes that degrade higher molecular weight DOM such as enzymes for aromatic carbon degradation, oxygenation, and decarboxylation, and (ii) releasing low molecular weight compounds and inorganic nutrients from DOM. However, growth stimulation following sunlight exposure of DOM came at a cost. Sunlight depleted the pool of aromatic compounds that supported microbial growth in the dark treatment, ultimately causing slower growth in the light treatment over 5 days. These first measurements of microbial metatranscriptomic responses to photo-alteration of DOM provide a mechanistic explanation for how sunlight exposure of terrigenous DOM alters microbial processing and respiration of DOM.
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[Evaluation of drug-drug interactions between yimitasvir phosphate capsules with sofosbuvir tablets, omeprazole magnesium enteric-coated tablets, and rosuvastatin calcium tablets].
Mai, JJ, Zhang, H, Peng, YY, Yang, X, Mao, L, Luo, L, Xie, HM, Zhang, YJ, Li, XJ, Ding, YH
Zhonghua gan zang bing za zhi = Zhonghua ganzangbing zazhi = Chinese journal of hepatology. 2020;(10):838-843
Abstract
Objective: To evaluate the drug-drug interactions and the tolerability of combined medication between yimitasvir phosphate capsules with sofosbuvir tablets, omeprazole magnesium enteric-coated tablets, and rosuvastatin calcium tablets in healthy volunteers. Methods: A randomized, open, and continuous administration design was used in trial 1 (yimitasvir phosphate capsules with sofosbuvir tablets). 28 subjects were randomly divided into two groups. A non-randomized, open design was used in trial 2 (yimitasvir phosphate capsules with omeprazole magnesium enteric-coated tablets), and included 42 subjects divided into three groups. The open design method was used in trial 3 (yimitasvir phosphate capsules with rosuvastatin calcium tablets), and included 14 subjects. The plasma concentrations of yimitasvir phosphate, sofosbuvir and their main metabolites GS-331007, omeprazole and rosuvastatin were validated by a liquid chromatography/tandem mass spectrometry (LC-MS/MS). The pharmacokinetic parameters were calculated by Phoenix winNonlin software. Results: (1) in trial 1, after single and co-administration, the 90% CI of sofosbuvir C(max) and AUC(0-tau) geometric mean ratio (GMR) were 152.0% (118.0% ~ 197.0%) and 230.0% (184.0% ~ 287.0%), with an increase of 52.0% and 130.0% compared to single dose of sofosbuvir, respectively. The 90% CI of GS-331007 C(max) GMR was 74.0% (67.5% ~ 81.2%) and reduced by 26% compared to single dose of sofosbuvir. (2) in trial 2, the 90% CI of C(max) GMR after yimitasvir single or co-administration at the same time, with a 4-hours interval, or with a 12- hours interval were 68.9% (44.5% ~ 106.7%) , 64.0% (43.8% ~ 93.6%) and 56.4%(38.9% ~ 81.9%), and the 90% CI of AUC(0-t) GMR were 68.6% (46.5% ~ 101.2%), 68.3% (47.6% ~ 98.0%) and 60.5% (41.8% ~ 87.5%), respectively. Compared with single dose of yimitasvir, the C(max) and AUC(0-t) were decreased by 31.1% and 31.4%, 36.0% and 31.7%, 43.6% and 39.5%, respectively. (3) In trial 3, after single and co-administration, the 90% CI of rosuvastatin C(max) and AUC(0-72) GMR were 172.4% (153.6% ~ 193.5%) and 158.0% (144.3% ~ 172.9%), respectively, with an increase of 74.9% and 60.5% compared to single dose of rosuvastatin. There were no serious adverse events and adverse events leading to withdrawal from the trial. Conclusion: Yimitasvir phosphate capsules have drug-drug interactions with sofosbuvir tablets, omeprazole magnesium enteric-coated tablets, and rosuvastatin calcium tablets.
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Experimental Exposure Assessment of Ionizable Organic Chemicals in In Vitro Cell-Based Bioassays.
Huchthausen, J, Mühlenbrink, M, König, M, Escher, BI, Henneberger, L
Chemical research in toxicology. 2020;(7):1845-1854
Abstract
Exposure assessment in in vitro cell-based bioassays is challenging for ionizable organic chemicals (IOCs), because they are present as more than one chemical species in the bioassay medium. Furthermore, compared to neutral organic chemicals, their binding to medium proteins and lipids is driven by more complex molecular interactions. Total medium concentrations (Ctotal,medium) and/or freely dissolved medium concentrations (Cfree,medium) were determined for one neutral chemical and 14 IOCs (acids, bases, multifunctional) at concentrations relevant for determination of cytotoxicity and effect. Cfree,medium was measured in two in vitro bioassays at the time of dosing and after 24 h of incubation using solid-phase microextraction. Cfree,medium was maximally 1.7 times lower than the nominal concentrations (Cnom) for the hydrophilic chemicals (caffeine and lamotrigine). For the organic acids (naproxen, ibuprofen, warfarin, and diclofenac), Cfree,medium was by a factor of 4 lower than Cnom at high concentrations, but the ratio was much higher at low concentrations, indicating a nonlinear binding behavior. The experimental Cfree,medium was also compared with Cfree,medium predicted with a mass balance model accounting for binding to medium proteins and lipids. The mass balance model performed well for five of the test chemicals (within a factor of 10), but it underestimated Cfree,medium by up to a factor of 1200 for chemicals that showed nonlinear binding to medium components. These findings emphasize that experimental exposure assessment is required for improved understanding of in vitro toxicity data.
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Metabolic engineering of microorganisms for production of aromatic compounds.
Huccetogullari, D, Luo, ZW, Lee, SY
Microbial cell factories. 2019;(1):41
Abstract
Metabolic engineering has been enabling development of high performance microbial strains for the efficient production of natural and non-natural compounds from renewable non-food biomass. Even though microbial production of various chemicals has successfully been conducted and commercialized, there are still numerous chemicals and materials that await their efficient bio-based production. Aromatic chemicals, which are typically derived from benzene, toluene and xylene in petroleum industry, have been used in large amounts in various industries. Over the last three decades, many metabolically engineered microorganisms have been developed for the bio-based production of aromatic chemicals, many of which are derived from aromatic amino acid pathways. This review highlights the latest metabolic engineering strategies and tools applied to the biosynthesis of aromatic chemicals, many derived from shikimate and aromatic amino acids, including L-phenylalanine, L-tyrosine and L-tryptophan. It is expected that more and more engineered microorganisms capable of efficiently producing aromatic chemicals will be developed toward their industrial-scale production from renewable biomass.
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Bacteria from the endosphere and rhizosphere of Quercus spp. use mainly cell wall-associated enzymes to decompose organic matter.
Lasa, AV, Mašínová, T, Baldrian, P, Fernández-López, M
PloS one. 2019;(3):e0214422
Abstract
Due to the ability of soil bacteria to solubilize minerals, fix N2 and mobilize nutrients entrapped in the organic matter, their role in nutrient turnover and plant fitness is of high relevance in forest ecosystems. Although several authors have already studied the organic matter decomposing enzymes produced by soil and plant root-interacting bacteria, most of the works did not account for the activity of cell wall-attached enzymes. Therefore, the enzyme deployment strategy of three bacterial collections (genera Luteibacter, Pseudomonas and Arthrobacter) associated with Quercus spp. roots was investigated by exploring both cell-bound and freely-released hydrolytic enzymes. We also studied the potential of these bacterial collections to produce enzymes involved in the transformation of plant and fungal biomass. Remarkably, the cell-associated enzymes accounted for the vast majority of the total activity detected among Luteibacter strains, suggesting that they could have developed a strategy to maintain the decomposition products in their vicinity, and therefore to reduce the diffusional losses of the products. The spectrum of the enzymes synthesized and the titres of activity were diverse among the three bacterial genera. While cellulolytic and hemicellulolytic enzymes were rather common among Luteibacter and Pseudomonas strains and less detected in Arthrobacter collection, the activity of lipase was widespread among all the tested strains. Our results indicate that a large fraction of the extracellular enzymatic activity is due to cell wall-attached enzymes for some bacteria, and that Quercus spp. root bacteria could contribute at different levels to carbon (C), phosphorus (P) and nitrogen (N) cycles.