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1.
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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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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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.
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The Synthesis of Organic Oils Blended Magnetorheological Fluids with the Field-Dependent Material Characterization.
Jinaga, R, Jagadeesha, T, Kolekar, S, Choi, SB
International journal of molecular sciences. 2019;(22)
Abstract
Automation is one of the trending terminologies in the field of engineering to achieve various sensors and actuators such as the hydraulic system. Smart fluid is also one of the hot topics for researchers to develop a type of actuator in many control systems since the fluid's rheological characteristics can be controlled or tuned by the intensity of the external stimuli. In this work, a new smart fluid of magnetorheological (MR) fluid is proposed and its field-dependent rheological characteristics are experimentally identified. An MR fluid using the carrier fluid as the blend of three different fluids, namely silicon oil, honey, and organic oil is prepared. In addition, two types of natural oils are used, sunflower oil and cottonseed oil. The samples are prepared using the blend as the carrier fluid, electrolytic iron powder coated with guar gum as the dispersed phase, and oleic acid as an additive. The quantity of oleic acid is optimized for 30% by weight of electrolytic iron powder. Two samples based on sunflower and cottonseed oil are synthesized and characterized for shear viscosity and shear stress with respect to shear rate subjected to a variable magnetic field. The blend-based MR fluid shows about 10% improvement over the sedimentation rate of silicon oil-based MR fluid as compared to that to conventional MR fluid. The cottonseed oil blend-based MR fluid performs better than sunflower-based fluid in terms of the viscosity and structure.
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Nitrate transformation and immobilization in particulate organic matter incubations: Influence of redox, iron and (a)biotic conditions.
Kizewski, FR, Kaye, JP, Martínez, CE
PloS one. 2019;(7):e0218752
Abstract
Nitrate can be reduced to other N inorganic species via denitrification and incorporated into organic matter by immobilization; however, the effect of biotic/abiotic and redox condition on immobilization and denitrification processes from a single system are not well documented. We hypothesize nitrate (NO3-) transformation pathways leading to the formation of dissolved- and solid-phase organic N are predominantly controlled by abiotic reactions, but the formation of soluble inorganic N species is controlled by redox condition. In this study, organic matter in the form of leaf compost (LC) was spiked with 15NO3- and incubated under oxic/anoxic and biotic/abiotic conditions at pH 6.5. We seek to understand how variations in environmental conditions impact NO3- transformation pathways through laboratory incubations. We find production of NH4+ is predominantly controlled by redox whereas NO3- conversion to dissolved organic nitrogen (DON) and immobilization in solid-phase N are predominantly controlled by abiotic processes. Twenty % of added 15N-NO3- was incorporated into DON under oxic conditions, with abiotic processes accounting for 85% of the overall incorporation. Nitrogen immobilization processes resulted in N concentrations of 4.1-6.6 μg N (g leaf compost)-1, with abiotic processes accounting for 100% and 66% of the overall (biotic+abiotic) N immobilization under anoxic and oxic conditions, respectively. 15N-NMR spectroscopy suggests 15NO3- was immobilized into amide/aminoquinones and nitro/oxime under anoxic conditions. A fraction of the NH4+ was produced abiotically under anoxic conditions (~10% of the total NH4+ production) although biotic organic N mineralization contributed to most of NH4+ production. Our results also indicate Fe(II) did not act as an electron source in biotic-oxic incubations; however, Fe(II) provided electrons for NO3- reduction in biotic-anoxic incubations although it was not the sole electron source. It is clear that, under the experimental conditions of this investigation, abiotic and redox processes play important roles in NO3- transformations. As climatic conditions change (e.g., frequency/intensity of rainfall), abiotic reactions that shift transformation pathways and N species concentrations from those controlled by biota might become more prevalent.
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Microwave-Assisted Extraction of Multiple Trace Levels of Intermediate Metabolites for Camptothecin Biosynthesis in Camptotheca acuminata and Their Simultaneous Determination by HPLC-LTQ-Orbitrap-MS/MS and HPLC-TSQ-MS.
Jin, Z, Wan, R, Yan, R, Su, Y, Huang, H, Zi, L, Yu, F
Molecules (Basel, Switzerland). 2019;(4)
Abstract
Camptothecin (CPT) has strong antitumor activity and is used as an anticancer therapeutic agent. To better understand and decipher the pathway of CPT biosynthesis in Camptotheca acuminata, the main purpose here was focused on creating an effective extraction strategy for a rich intermediate metabolite profile. In the present study, a 70% aqueous acetonitrile was verified as an optimal extraction solvent for microwave-assisted extraction (MAE) of metabolites by spiking experiments. Based on multi-objective optimization, the best extraction conditions of a solid-liquid ratio of 1:20, microwave power of 230 W, and a time of 4 min were achieved using a full factorial 3⁴ experimental design. Crude extracts obtained from the shoot apex of C. acuminata using MAE have been qualitatively profiled by high-performance liquid chromatography coupled with linear ion trap quadrupole-orbitrap mass spectrometry (HPLC-LTQ-Orbitrap-MS/MS) and a HPLC triple quadrupole-MS (HPLC-TSQ-MS) analysis was conducted for their metabolite content in different tissues. CPT, and ten related metabolites and their isomers, including tryptamine, loganic acid, secologanic acid, strictosidinic acid, strictosamide, strictosamide epoxide, strictosamide diol, strictosamide ketolactam, pumiloside, and deoxypumiloside, were detected and tentatively identified. Scanning electron microscopy (SEM) imaging of the shoot apex demonstrated that severe cell disruption was evident after intensified extraction processes. The study showed the difference of metabolite profiles and the enhancement of metabolite content after microwave-pretreated techniques, and the established MAE procedure is an effective methodology to preserve valuable metabolite compounds for analysis.
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10.
Modulating Inflammation in Monocytes Using Capillary Fiber Organic Electronic Ion Pumps.
Seitanidou, M, Blomgran, R, Pushpamithran, G, Berggren, M, Simon, DT
Advanced healthcare materials. 2019;(19):e1900813
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Abstract
An organic electronic ion pump (OEIP) delivers ions and drugs from a source, through a charge selective membrane, to a target upon an electric bias. Miniaturization of this technology is crucial and will provide several advantages, ranging from better spatiotemporal control of delivery to reduced invasiveness for implanted OEIPs. To miniaturize OEIPs, new configurations have been developed based on glass capillary fibers filled with an anion exchange membrane (AEM). Fiber capillary OEIPs can be easily implanted in proximity to targeted cells and tissues. Herein, the efficacy of such a fiber capillary OEIP for modulation of inflammation in human monocytes is demonstrated. The devices are located on inflammatory monocytes and local delivery of salicylic acid (SA) is initiated. Highly localized SA delivery results in a significant decrease in cytokine (tumor necrosis factor alpha and interleukin 6) levels after lipopolysaccharide stimulation. The findings-the first use of such capillary OEIPs in mammalian cells or systems-demonstrate the utility of the technology for optimizing transport and delivery of different therapeutic substances at low concentrations, with the benefit of local and controlled administration that limits the adverse effect of oral/systemic drug delivery.