Broad Tolerance Research Articles

Polyhydroxyalkanoate Copolymer Production by Recombinant Ralstonia eutropha Strain 1F2 from Fructose or Carbon Dioxide as Sole Carbon Source.

Ralstonia eutropha strain H16 is a chemoautotrophic bacterium that oxidizes hydrogen and accumulates poly[(R)-3-hydroxybutyrate] [P(3HB)], a prominent polyhydroxyalkanoate (PHA), within its cell. R. eutropha utilizes fructose or CO2 as its sole carbon source for this process. A PHA-negative mutant of strain H16, known as R. eutropha strain PHB-4, cannot produce PHA. Strain 1F2, derived from strain PHB-4, is a leucine analog-resistant mutant. Remarkably, the recombinant 1F2 strain exhibits the capacity to synthesize 3HB-based PHA copolymers containing 3-hydroxyvalerate (3HV) and 3-hydroxy-4-methyvalerate (3H4MV) comonomer units from fructose or CO2. This ability is conferred by the expression of a broad substrate-specific PHA synthase and tolerance to feedback inhibition of branched amino acids. However, the total amount of comonomer units incorporated into PHA was up to around 5 mol%. In this study, strain 1F2 underwent genetic engineering to augment the comonomer supply incorporated into PHA. This enhancement involved several modifications, including the additional expression of the broad substrate-specific 3-ketothiolase gene (bktB), the heterologous expression of the 2-ketoacid decarboxylase gene (kivd), and the phenylacetaldehyde dehydrogenase gene (padA). Furthermore, the genome of strain 1F2 was altered through the deletion of the 3-hydroxyacyl-CoA dehydrogenase gene (hbdH). The introduction of bktB-kivd-padA resulted in increased 3HV incorporation, reaching 13.9 mol% from fructose and 6.4 mol% from CO2. Additionally, the hbdH deletion resulted in the production of PHA copolymers containing (S)-3-hydroxy-2-methylpropionate (3H2MP). Interestingly, hbdH deletion increased the weight-average molecular weight of the PHA to over 3.0 × 106 on fructose. Thus, it demonstrates the positive effects of hbdH deletion on the copolymer composition and molecular weight of PHA.

Visible Light-Catalyzed Reactions of Polysulfide (DBSPS) with Aryldiazonium.

A visible light-catalyzed radical coupling reaction of polysulfide reagents with aryldiazonium was developed, which gave thiosulfonates under mild conditions. In this reaction, the thiosulfonates were isolated in good yields with a broad tolerance to functional groups. And the synthesis of diaryl monosulfides were achieved through a step-by-step reaction of two molecular aryldiazonium with DBSPS, where the sulfur source was provided by DBSPS. It was worth noting that the reaction of this monosulfides could also be achieved by a one pot two-step process. The described polysulfide reagents were able to produce three new radicals: sulfonyl radicals, sulfur-sulfonyl radicals and sulfur-sulfur-sulfonyl radicals.

Stereodivergent Synthesis of Spiroaminals via Chiral Bifunctional Hydrogen Bonding Organocatalysis

AbstractSpiroaminals represent novel structural motifs prevalent in diverse natural products and biologically active molecules. Achieving their enantioselective synthesis is a highly desirable and challenging task in synthetic endeavors due to their intricate molecular frameworks. Herein, we accomplished the first stereodivergent construction of spiroaminals using chiral bifunctional organocatalyzed intramolecular 1,2‐addition followed by an oxa‐Michael addition cascade in a high atom and step economical pathway. A proper modulation of the cinchona‐derived squaramide catalysts efficiently provided access to all the possible stereoisomers with high yield, diastereoselectivity, and excellent enantioselectivity while displaying a broad substrate tolerance. Additionally, we validated the scalability of the reaction and demonstrated the synthesis of variable spiroaminal scaffolds, confirming the viability of our protocol.

Stereodivergent Synthesis of Spiroaminals via Chiral Bifunctional Hydrogen Bonding Organocatalysis.

Spiroaminals represent novel structural motifs prevalent in diverse natural products and biologically active molecules. Achieving their enantioselective synthesis is a highly desirable and challenging task in synthetic endeavors due to their intricate molecular frameworks. Herein, we accomplished the first stereodivergent construction of spiroaminals using chiral bifunctional organocatalyzed intramolecular 1,2-addition followed by an oxa-Michael addition cascade in a high atom and step economical pathway. A proper modulation of the cinchona-derived squaramide catalysts efficiently provided access to all the possible stereoisomers with high yield, diastereoselectivity, and excellent enantioselectivity while displaying a broad substrate tolerance. Additionally, we validated the scalability of the reaction and demonstrated the synthesis of variable spiroaminal scaffolds, confirming the viability of our protocol.

H2 production from coal by enriching sugar fermentation and alkane oxidation with hyperthermophilic resistance microbes in municipal wastewater

Anaerobic hydrogen production from coal provides an alternative to achieve clean energy and zero greenhouse gas emissions. However, the recalcitrant structure of coal is the barrier for microorganisms to access, leading to inefficient hydrogen production. Municipal wastewater contains rich microbial resources with high degradability and a broad tolerance range to temperature, which was herein used to drive hydrogen production from coal at 35, 55 and 65 °C. The result showed the total hydrogen production (1.02 mL/g TS) at 65 °C was 34.0 and 51.0 times the 35 °C and 55 °C, respectively. For 65 °C, the decomposability of C-H linkage and aromatic ring structures in coal was the best; sugar fermentation and alkane oxidation pathways were effectively enriched, leading to the highest concentration of volatile fatty acids and alkanes. Which resulted in a shift of dominant bacteria from the lactate- and hydrogen-producing bacterium Bacillus at 35 °C and 55 °C to the hydrogen-producing bacterium Thermoanaerobacterium at 65 °C. Moreover, a gradient temperature cultivation from 35 to 65 °C (gradient increase temperature, GIT) based on the domestication concept was conducted. It is surprising that the GIT group displayed significantly lower hydrogen production (0.14 mL/g TS) compared to 65 °C group, because bacteria can adapt from mesophilic to hyperthermophilic temperatures, including Bacillus, Fonticella, Lysinibacillus, Ureibacillus, Microbacterium, and Geobacillus, inhibiting the Thermoanaerobacterium in the GIT group by producing lactic acid and competing for living space. This study proposes an alternative for efficient hydrogen production from coal. In the future, hydrogen production can be further enhanced through both substrate pretreatment and inoculum domestication, the related core mechanism will also be focus on.

Cesium Carbonate Promoted Direct Amidation of Unactivated Esters with Amino Alcohol Derivatives.

Cesium carbonate promoted direct amidation of unactivated esters with amino alcohols was developed without the use of transition-metal catalysts and coupling reagents. This method enabled the synthesis of several serine-containing oligopeptides and benzamide derivatives with yields up to 90%. The methodology proceeds under mild reaction conditions and exhibits no racemization for most naturally occurring amino acid substrates. The reaction demonstrates good compatibility with primary alkyl and benzyl esters and broad tolerance for a range of amino acid substrates with nonpolar and protected side chains. The hydroxy group on the amine nucleophile was found to be critical for the reaction to be successful. A likely mechanism involving cesium coordination to the substrates enabling the subsequent proximity-driven acyl transfer was proposed. The practicality of this approach was demonstrated in the preparation of a biologically active nicotinamide derivative in a reasonable yield.

Tag-free protein modification by lipoate ligase A: exploring substrate tolerance.

This study delves into the functional intricacies of lipoate ligase A (LplA), an enzyme showing great promise in bioconjugation due to its unique capacity for introducing azido groups into proteins without requiring a genetic tag. We aimed to enhance the understanding of LplA's functionality, particularly its substrate tolerance and the reliability of various analytical techniques. A pivotal aspect of our approach was incorporating azido groups into a range of proteins, followed by the addition of the fluorescent molecule Cy3 via click chemistry. Analysis of fluorescent intensity in the altered proteins indicated varying degrees of conjugation. Additionally, phenyl resin-based RP-HPLC facilitated effective separation of modified proteins, unmodified proteins, and remaining fluorescent tags post-separation. SASA analysis provided insights into conjugation trends, guiding the identification of proteins amenable to LplA's tag-free modification. Our findings demonstrate LplA's broad substrate tolerability for protein modification.

Photoinduced rapid Chloropropylation of Quinoxalin-2(1H)-ones with Cyclopropane and HCl

Herein, a photoinduced three-component reaction of aryl cyclopropane, quinoxalin-2(1H)-ones and HCl has been developed to introduce chloroalkyl motifs into N-heteroarenes. This synthetic method provides a green and efficient way to realize the chloropropylation of quinoxalin-2(1H)-ones and afford a series of chlorinated quinoxalin-2(1H)-one derivatives in moderate to good yields with the broad functional-group tolerance.

Cyclic Amine Synthesis via Catalytic Radical-Polar Crossover Cycloadditions.

The rapid assembly of valuable cyclic amine architectures in a single step from simple precursors has been recognized as an ideal platform in term of efficiency and sustainability. Although a vast number of studies regarding cyclic amine synthesis has been reported, new synthetic disconnection approaches are still high in demand. Herein, we report a catalytic radical-polar crossover cycloaddition to cyclic amine synthesis triggered from primary sulfonamide under photoredox condition. This newly developed disconnection, comparable to established synthetic approaches, will allow to construct β, β-disubstituted cyclic amine and β-monosubstituted cyclic amine derivatives efficiently. This study highlights the unique utility of primary sulfonamide as a bifunctional reagent, which acts as a radical precursor and a nucleophile. The open-shell methodology demonstrates broad tolerance to various functional groups, drug derivatives and natural products in an economically and sustainable fashion.

Cyclic Amine Synthesis via Catalytic Radical‐Polar Crossover Cycloadditions

AbstractThe rapid assembly of valuable cyclic amine architectures in a single step from simple precursors has been recognized as an ideal platform in term of efficiency and sustainability. Although a vast number of studies regarding cyclic amine synthesis has been reported, new synthetic disconnection approaches are still high in demand. Herein, we report a catalytic radical‐polar crossover cycloaddition to cyclic amine synthesis triggered from primary sulfonamide under photoredox condition. This newly developed disconnection, comparable to established synthetic approaches, will allow to construct β, β‐disubstituted cyclic amine and β‐monosubstituted cyclic amine derivatives efficiently. This study highlights the unique utility of primary sulfonamide as a bifunctional reagent, which acts as a radical precursor and a nucleophile. The open‐shell methodology demonstrates broad tolerance to various functional groups, drug derivatives and natural products in an economically and sustainable fashion.

Unravelling the biogeographic determinants of tree growth sensitivity to freeze and drought in Canada's forests

Abstract The evolutionary dynamics of tree species are influenced by their specific climatic environments, and their ability to persist is determined by adaptive strategies such as broad climate tolerance, phenotypic plasticity or genetic differentiation. Biogeographical predictions indicate that populations located at the edges of their ranges are more likely to experience heightened vulnerability to climatic fluctuations due to approaching tolerance limits. However, if local phenotypic acclimation or genetic adaptation has taken place, trees near the edge of their range could demonstrate comparable sensitivity to freeze and drought as the rest of the sampled population. Nevertheless, there remains uncertainty regarding the extent of developmental and evolutionary adjustments in climate sensitivity across the entire ranges of many tree species. Here, we document the biogeography of tree growth sensitivity to freeze and soil water and vapour pressure deficits during 1950–2018 using an extensive multi‐species tree‐ring dataset of 35,784 trees at 4535 sites covering boreal, temperate conifer and temperate deciduous forests of Canada. We quantify the relationships between tree radial growth increment and seasonal climate variables and explore factors driving the observed patterns of annual growth and climate sensitivity such as species, regional climate and local site conditions, and tree age and size. Freeze and drought had widespread impacts on tree growth that were contingent on the presence of focal species in interaction with tree size. An indirect growth thermal limitation towards cold/wet edges, occurring through the site moisture conditions, was observed in seven widespread species (e.g. Picea glauca, Picea mariana and Larix laricina). Moreover, six species had negative drought impacts more strongly expressed towards their warm/dry edges (e.g. Abies balsamea, Betula papyrifera and Pseudotsuga menziesii). However, widespread Picea, Pinus and Populus species showed no indication of increased sensitivity to soil water conditions at these edges. Synthesis. Our findings support the idea of evolutionary or acclimatization adjustments in the development of populations in response to long‐term climate conditions experienced in their respective locations. This underscores the importance of incorporating phenotypic and genomic data into future analyses of climate change impacts, which would enhance our ability to predict potential ecological shifts.

Niche appropriation by a widespread nereidid polychaete within estuarine-like modern coastal stromatolite pools

Determining how organisms respond to environmental gradients or variability is a central theme in ecology, which can be used to predict niche occupation. In this study we assessed the natural population variability of a locally widespread nereidid Ceratonereis (Composetia) keiskama to determine whether its broad distribution can be used to infer a generalist autecological niche. This was examined in the recently discovered estuarine-like stromatolite-forming pools in South Africa where this polychaete also occurs. Results suggest that neither temperature nor salinity were important predictors of C. keiskama density, attesting to its broad ecophysiological tolerance. Instead, sediment properties were correlated with presence in estuaries and the stromatolite pools, which suggests that foraging conditions or biotic interactions are more important autecological drivers. Notably, C. keiskama density in the stromatolite pools was on average an order of magnitude higher than in most estuaries and this species is the dominant infaunal and epibenthic macroinvertebrate within the stromatolite sediment, suggesting an opportunistic occupation of this niche. These findings highlight the importance of linking species ranges to niche occupation of diverse habitat types, and it frames future work which should address the population level connectivity and gene transfer between such different environments.

Complete mitochondrial genome of the Starhead Topminnow Fundulus dispar (Cyprinodontiformes: Fundulidae)

Topminnows of the Teleost genus Fundulus serve as model organisms in ecotoxicology because of their broad physiological tolerance and propensity to breed in captivity. This research has been primarily limited to intraspecific comparisons, due to incomplete understanding of the evolutionary history of the genus, which is necessary for use of phylogenetic comparative methods. Interspecific relationships of topminnows remain unresolved, despite recent advances in mitochondrial and nuclear genome sequencing. Specifically, interrelationships of a group containing the starhead topminnows (Fundulus blairae, F. dispar, F. escambiae, F. lineolatus, and F. nottii) typically yield low node support values. Here, we present the first annotated mitochondrial genome of the Starhead Topminnow (F. dispar) and provide a phylogenetic hypothesis for starhead topminnows within the genus Fundulus. DNA was isolated from a specimen of F. dispar collected in Kentucky, USA. The circular genome is 16,564 bp long and contains 13 protein-coding genes, two ribosomal RNAs (rRNA), 22 transfer RNAs (tRNA), and one control region (D-loop). Our phylogenetic analysis supports a sister relationship between F. dispar and a group containing F. notatus and F. olivaceus. This data helps to resolve the phylogenetic placement of starhead topminnows.

Silver-catalyzed direct conversion of epoxides into cyclopropanes using N-triftosylhydrazones

Epoxides, as a prominent small ring O-heterocyclic and the privileged pharmacophores for medicinal chemistry, have recently represented an ideal substrate for the development of single-atom replacements. The previous O-to-C replacement strategy for epoxides to date typically requires high temperatures to achieve low yields and lacks substrate range and functional group tolerance, so achieving this oxygen-carbon exchange remains a formidable challenge. Here, we report a silver-catalyzed direct conversion of epoxides into trifluoromethylcyclopropanes in a single step using trifluoromethyl N-triftosylhydrazones as carbene precursors, thereby achieving oxygen-carbon exchange via a tandem deoxygenation/[2 + 1] cycloaddition. The reaction shows broad tolerance of functional groups, allowing routine cheletropic olefin synthesis in a strategy for the net oxygen-carbon exchange reaction. The utility of this method is further showcased with the late-stage diversification of epoxides derived from bioactive natural products and drugs. Mechanistic experiments and DFT calculations elucidate the reaction mechanism and the origin of the chemo- and stereoselectivity.

Physiological Responses of C4 Perennial Bioenergy Grasses to Climate Change: Causes, Consequences, and Constraints.

C4 perennial bioenergy grasses are an economically and ecologically important group whose responses to climate change will be important to the future bioeconomy. These grasses are highly productive and frequently possess large geographic ranges and broad environmental tolerances, which may contribute to the evolution of ecotypes that differ in physiological acclimation capacity and the evolution of distinct functional strategies. C4 perennial bioenergy grasses are predicted to thrive under climate change-C4 photosynthesis likely evolved to enhance photosynthetic efficiency under stressful conditions of low [CO2], high temperature, and drought-although few studies have examined how these species will respond to combined stresses or to extremes of temperature and precipitation. Important targets for C4 perennial bioenergy production in a changing world, such as sustainability and resilience, can benefit from combining knowledge of C4 physiology with recent advances in crop improvement, especially genomic selection.

Oxygen-vacancies rich CuFe2O4 catalyst as efficient peroxymonosulfate activator for enhanced oxytetracycline degradation: Performance and mechanism

Peroxymonosulfate (PMS) based advanced oxidation methods have gained attention for eliminating antibiotics like oxytetracycline (OTC) from wastewater. This study presents an innovative approach by incorporating oxygen vacancies (OVs) into CuFe2O4 to create OVs-CuFe2O4 materials. Compared to traditional CuFe2O4, OVs-CuFe2O4 offered superior catalytic performance due to its surface-rich OVs promoting rapid conversion of oxygen and efficient metal redox of Fe(Ⅱ)/Fe(Ⅲ) and Cu(Ⅰ)/Cu(Ⅱ). OVs-CuFe2O4/PMS achieved impressive 95.7 % OTC removal in 45 min with a rate constant of 0.107 min−1, which was 3.87 times of CuFe2O4/PMS. The OVs-CuFe2O4/PMS system exhibited a broad pH tolerance range of 3 to 11 with OTC removal efficiency above 89.3 %. The OVs-CuFe2O4/PMS system produced various free radicals and non-radical for OTC degradation. The OVs-CuFe2O4 also has good stability with removal efficiency of 81.9 % at fourth cycle. This research gives a strategy to construct oxygen vacancies enrich catalysts for efficient activation of PMS and treatment of antibiotics wastewater.

Efficient bio-remediation of multiple aromatic hydrocarbons using different types of thermotolerant, ring-cleaving dioxygenases derived from Aeribacillus pallidus HB-1

As toxic contaminants, aromatic compounds are widespread in most environmental matrices, and bioenzymatic catalysis plays a critical role in the degradation of xenobiotics. Here, a thermophillic aromatic hydrocarbon degrader Aeribacillus pallidus HB-1 was found. Bioinformatic analysis of the HB-1 genome revealed two ring-cleaving extradiol dioxygenases (EDOs), among which, EDO-0418 was assigned to a new subfamily of type I.1 EDOs and exhibited a broad substrate specificity, particularly towards biarylic substrate. Both EDOs exhibited optimal activities at elevated temperatures (55 and 65 °C, respectively) and showed remarkable thermostability, pH stability, metal ion resistance and tolerance to chemical reagents. Most importantly, simulated wastewater bioreactor experiments demonstrated efficient and uniform degradation performance of mixed aromatic substrates under harsh environments by the two enzymes combined for potential industrial applications. The unveiling of two thermostable dioxygenases with broad substrate specificities and stress tolerance provides a novel approach for highly efficient environmental bioremediation using composite enzyme systems.

Salinity-mediated limitation of asexual reproduction in the colonial ascidian Polyandrocarpa zorritensis

Ascidians are among the most common invasive marine invertebrates worldwide. Many species of non-indigenous ascidians (NIAs) have successfully colonized the Mediterranean Sea, notably within anthropized coastal lagoons and harbors. Although invasive species are generally characterized by their broad ecological tolerance, different ascidian species exhibit varied responses to biotic and abiotic environmental stressors, including temperature and salinity. Acquiring a better understanding about of the impact of such parameters on ascidian life history is crucial for predicting the invasive potential of NIAs. In this study, we investigated the impact of various salinities on the reproduction of the colonial ascidian Polyandorcarpa zorritensis, a species indigenous to Peru and a thriving invader. P. zorritensis undergoes asexual reproduction via a peculiar form of budding named vasal budding and produces resistant spherules, which likely facilitated its dissemination over long distances. Despite its widespread distribution along the Pacific and Atlantic coasts, it is only found in a few Mediterranean coastal areas with a low salinity. We tested the impact of different salinity conditions on the sexual and asexual reproduction rates of P. zorritensis in a controlled laboratory setting. Our experiments showed that the rate of asexual reproduction in colonies bred at 29 or 36 ppt salinity levels, corresponding to the natural range inhabited by P. zorritensis, was higher than those grown in 40 ppt salinity, commonly found in Mediterranean marinas and harbors. The results suggest that, although P. zorritensis has been present in the Mediterranean for several decades, its potential for invasion could be constrained by an intolerance to high salinity.

Lignin bioconversion based on genome mining for ligninolytic genes in Erwinia billingiae QL-Z3

BackgroundBioconversion of plant biomass into biofuels and bio-products produces large amounts of lignin. The aromatic biopolymers need to be degraded before being converted into value-added bio-products. Microbes can be environment-friendly and efficiently degrade lignin. Compared to fungi, bacteria have some advantages in lignin degradation, including broad tolerance to pH, temperature, and oxygen and the toolkit for genetic manipulation.ResultsOur previous study isolated a novel ligninolytic bacterial strain Erwinia billingiae QL-Z3. Under optimized conditions, its rate of lignin degradation was 25.24% at 1.5 g/L lignin as the sole carbon source. Whole genome sequencing revealed 4556 genes in the genome of QL-Z3. Among 4428 protein-coding genes are 139 CAZyme genes, including 54 glycoside hydrolase (GH) and 16 auxiliary activity (AA) genes. In addition, 74 genes encoding extracellular enzymes are potentially involved in lignin degradation. Real-time PCR quantification demonstrated that the expression of potential ligninolytic genes were significantly induced by lignin. 8 knock-out mutants and complementary strains were constructed. Disruption of the gene for ELAC_205 (laccase) as well as EDYP_48 (Dyp-type peroxidase), ESOD_1236 (superoxide dismutase), EDIO_858 (dioxygenase), EMON_3330 (monooxygenase), or EMCAT_3587 (manganese catalase) significantly reduced the lignin-degrading activity of QL-Z3 by 47–69%. Heterologously expressed and purified enzymes further confirmed their role in lignin degradation. Fourier transform infrared spectroscopy (FTIR) results indicated that the lignin structure was damaged, the benzene ring structure and groups of macromolecules were opened, and the chemical bond was broken under the action of six enzymes encoded by genes. The abundant enzymatic metabolic products by EDYP_48, ELAC_205 and ESOD_1236 were systematically analyzed via liquid chromatography–mass spectrometry (LC–MS) analysis, and then provide a speculative pathway for lignin biodegradation. Finally, The activities of ligninolytic enzymes from fermentation supernatant, namely, LiP, MnP and Lac were 367.50 U/L, 839.50 U/L, and 219.00 U/L by orthogonal optimization.ConclusionsOur findings provide that QL-Z3 and its enzymes have the potential for industrial application and hold great promise for the bioconversion of lignin into bioproducts in lignin valorization.

Hydroxylation of Aryl Sulfonium Salts for Phenol Synthesis under Mild Reaction Conditions.

Hydroxylation of aryl sulfonium salts could be realized by utilizing acetohydroxamic acid and oxime as hydroxylative agents in the presence of cesium carbonate as a base, leading to a variety of structurally diverse hydroxylated arenes in 47-95% yields. In addition, the reaction exhibited broad functionality tolerance, and a range of important functional groups (e.g., cyano, nitro, sulfonyl, formyl, keto, and ester) could be well amenable to the mild reaction conditions.