Small molecule organocatalysts are abundant in all living organisms. However, their use as organocatalysts in cells has been underexplored. Herein, we report that organocatalytic aldol chemistry can be interfaced with living Escherichia coli to enable the α-methylenation of cellular aldehydes using biogenic amines such as L-Pro or phosphate. The biocompatible reaction is mild and can be interfaced with butyraldehyde generated from D-glucose via engineered metabolism to enable the production of 2-methylenebutanal (2-MB) and 2-methylbutanal (2-MBA) by anaerobic fermentation, and 2-methylbutanol (2-MBO) by whole-cell catalysis. Overall, this study demonstrates the combination of non-enzymatic organocatalytic and metabolic reactions in vivo for the sustainable synthesis of valuable non-natural chemicals that cannot be accessed using enzymatic chemistry alone. 相似文献
The introduction of new reactivity into living organisms is a major challenge in synthetic biology. Despite an increasing interest in both the development of small‐molecule catalysts that are compatible with aqueous media and the engineering of enzymes to perform new chemistry in vitro, the integration of non‐native reactivity into metabolic pathways for small‐molecule production has been underexplored. Herein we report a biocompatible iron(III) phthalocyanine catalyst capable of efficient olefin cyclopropanation in the presence of a living microorganism. By interfacing this catalyst with E. coli engineered to produce styrene, we synthesized non‐natural phenyl cyclopropanes directly from D ‐glucose in single‐vessel fermentations. This process is the first example of the combination of nonbiological carbene‐transfer reactivity with cellular metabolism for small‐molecule production. 相似文献
Microorganisms can be programmed to perform chemical synthesis via metabolic engineering. However, despite an increasing interest in the use of de novo metabolic pathways and designer whole‐cells for small molecule synthesis, the inherent synthetic capabilities of native microorganisms remain underexplored. Herein, we report the use of unmodified E. coli BL21(DE3) cells for the reduction of keto‐acrylic compounds and apply this whole‐cell biotransformation to the synthesis of aminolevulinic acid from a lignin‐derived feedstock. The reduction reaction is rapid, chemo‐, and enantioselective, occurs under mild conditions (37 °C, aqueous media), and requires no toxic transition metals or external reductants. This study demonstrates the remarkable promiscuity of central metabolism in bacterial cells and how these processes can be leveraged for synthetic chemistry without the need for genetic manipulation. 相似文献
Enzyme engineering is an indispensable tool in the field of synthetic biology, where enzymes are challenged to carry out novel or improved functions. Achieving these goals sometimes goes beyond modifying the primary sequence of the enzyme itself. The use of protein or nucleic acid scaffolds to enhance enzyme properties has been reported for applications such as microbial production of chemicals, biosensor development and bioremediation. Key advantages of using these assemblies include optimizing reaction conditions, improving metabolic flux and increasing enzyme stability. This review summarizes recent trends in utilizing genetically encodable scaffolds, developed in line with synthetic biology methodologies, to complement the purposeful deployment of enzymes. Current molecular tools for constructing these synthetic enzyme-scaffold systems are also highlighted. 相似文献
Organic chemists and metabolic engineers use orthogonal technologies to construct essential small molecules such as pharmaceuticals and commodity chemicals. While chemists have leveraged the unique capabilities of biological catalysts for small‐molecule production, metabolic engineers have not likewise integrated reactions from organic synthesis with the metabolism of living organisms. Reported herein is a method for alkene hydrogenation which utilizes a palladium catalyst and hydrogen gas generated directly by a living microorganism. This biocompatible transformation, which requires both catalyst and microbe, and can be used on a preparative scale, represents a new strategy for chemical synthesis that combines organic chemistry and metabolic engineering. 相似文献
A targeted micellar drug delivery system is developed from a biocompatible and biodegradable amphiphilic polyester, poly(Lac‐OCA)‐b‐(poly(Tyr(alkynyl)‐OCA)‐g‐mannose) (PLA‐b‐(PTA‐g‐mannose), that is synthesized via controlled ring‐opening polymerization of O‐carboxyanhydride (OCA) and highly efficient “Click” chemistry. Doxorubicin (DOX), a model lipophilic anticancer drug, can be effectively encapsulated into the micelles, and the mannose moiety allows active targeting of the micelles to cancer cells that specifically express mannose receptors, which thereafter enhances the anticancer efficiency of the drug. Comprised entirely of biodegradable and biocompatible polyesters, this micellar system demonstrates promising potentials for targeted drug delivery and cancer therapy.
Fluorine has become an important element for the design of synthetic molecules for use in medicine, agriculture, and materials. Despite the many advantages provided by fluorine for tuning key molecular properties, it is rarely found in natural metabolism. We seek to expand the molecular space available for discovery through the development of new biosynthetic strategies that cross synthetic with natural compounds. Towards this goal, we engineered a microbial host for organofluorine metabolism and show that we can achieve the production of the fluorinated diketide 2‐fluoro‐3‐hydroxybutyrate at approximately 50 % yield. This fluorinated diketide can be used as a monomer in vivo to produce fluorinated poly(hydroxyalkanoate) (PHA) bioplastics with fluorine substitutions ranging from around 5–15 %. This system provides a platform to produce mm flux through the key fluoromalonyl coenzyme A (CoA) building block, thereby offering the potential to generate a broad range of fluorinated small‐molecule targets in living cells. 相似文献
Poly-(3-hydroxybutyrate) (PHB) is a polyester with biodegradable and biocompatible characteristics and has many potential applications. To reduce the raw material costs and microbial energy consumption during PHB production, cheaper carbon sources such as sucrose were evaluated for the synthesis of PHB under anaerobic conditions. In this study, metabolic network analysis was conducted to construct an optimized pathway for PHB production using sucrose as the sole carbon source and to guide the gene knockout to reduce the generation of mixed acid byproducts. The plasmid pMCS-sacC was constructed to utilize sucrose as a sole carbon source, and the cascaded promoter P3nirB was used to enhance PHB synthesis under anaerobic conditions. The mixed acid fermentation pathway was knocked out in Escherichia coli S17-1 to reduce the synthesis of byproducts. As a result, PHB yield was improved to 80% in 6.21 g/L cell dry weight by the resulted recombinant Escherichia coli in a 5 L bed fermentation, using sucrose as the sole carbon source under anaerobic conditions. As a result, the production costs of PHB will be significantly reduced. 相似文献
Engineering enzymes with novel reactivity and applying them in metabolic pathways to produce valuable products are quite challenging due to the intrinsic complexity of metabolic networks and the need for high in vivo catalytic efficiency. Triacetic acid lactone (TAL), naturally generated by 2-pyrone synthase (2PS), is a platform molecule that can be produced via microbial fermentation and further converted into value-added products. However, these conversions require extra synthetic steps under harsh conditions. We herein report a biocatalytic system for direct generation of TAL derivatives under mild conditions with controlled chemoselectivity by rationally engineering the 2PS active site and then rewiring the biocatalytic pathway in the metabolic network of E. coli to produce high-value products, such as kavalactone precursors, with yields up to 17 mg/L culture. Computer modeling indicates sterics and hydrogen-bond interactions play key roles in tuning the selectivity, efficiency and yield. 相似文献
A microarray enables high-throughput interaction screening of numerous biomolecules; however, fabrication of a microarray composed of cellular membrane components has proven difficult. We report fabrication of a liposomal glyco-microarray by using an azide-reactive liposome that carries synthetic and natural glycolipids via chemically selective and biocompatible liposome immobilization chemistry. Briefly, liposomes carrying anchor lipid dipalmitoylphosphatidylethanolamine (DPPE)-PEG(2000)-triphenylphosphine and ganglioside (GM1 or GM3) were prepared first and were then printed onto an azide-modified glass slide so as to afford a liposomal glyco-microarray via Staudinger ligation. Fluorescent dye release kinetics and fluorescence imaging confirmed successful liposome immobilization and specific protein binding to the intact arrayed glycoliposomes. The liposomal glyco-microarray with different gangliosides showed their specific lectin and toxin binding with different binding affinity. The azide-reactive liposome provides a facile strategy for fabrication of either a natural or a synthetic glycolipid-based membrane-mimetic glycoarray. This liposomal glyco-microarray is simple and broadly applicable and thus will find important biomedical applications, such as studying glycolipid-protein interactions and toxin screening applications. 相似文献
The self‐assembled nanostructures of a high‐molecular‐weight rod–coil block copolymer, poly(styrene‐block‐(2,5‐bis[4‐methoxyphenyl]oxycarbonyl)styrene) (PS‐b‐PMPCS), in p‐xylene are studied. The cylindrical micelles, long segmental cylindrical micelle associates, spherical micelles, and spherical micelle associates are observed with increased copolymer concentration. The high molecular weight of PS leads to the entanglement between PS chains from different micelles, which is the force for supramolecular interactions. Short cylindrical micelles are connected end‐to‐end via this supramolecular chemistry to form long segmental cylindrical micelle associates, analogue to the condensation polymerization process, with direction and saturation. On the other hand, spherical micelles assemble via supramolecular chemistry to form spherical micelle associates, yet without any direction due to their isotropic properties.
P‐113, AKRHHGYKRKFH‐NH2, was derived from human saliva and found to possess clinical activity against fungus infections in HIV patients with oral candidiasis. We have determined the solution structure of P‐113 bound to membrane‐mimetic SDS micelles by two‐dimensional NMR methods. The SDS micelle‐bound structure of P‐113 adopts an α‐helical segment and the positively charged residues are clustered together to form a hydrophilic patch. A variety of biophysical and biochemical experiments, including circular dichroism, fluorescence spectroscopy and microcalorimetry, were used to show that P‐113 interacted with negatively charged phospholipid vesicles and induced dye release from these vesicles. However, its dye leakage efficiency is much less than the results of previously reported antimicrobial peptides. These results suggest that the antimicrobial activity of P‐113, unlike other antimicrobial peptides, may act not only through binding to and destabilization of the microbial membrane but also through a specific protein receptor on the microbial cell surface. 相似文献
The aim of this work was to further investigate the glycolysis performance of lager and ale brewer??s yeasts under different fermentation temperature using a combined analysis of metabolic flux, glycolytic enzyme activities, and flux control. The results indicated that the fluxes through glycolytic pathway decreased with the change of the fermentation temperature from 15?°C to 10?°C, which resulted in the prolonged fermentation times. The maximum activities (Vmax) of hexokinase (HK), phosphofructokinase (PFK), and pyruvate kinase (PK) at key nodes of glycolytic pathway decreased with decreasing fermentation temperature, which was estimated to have different control extent (22?C84?%) on the glycolytic fluxes in exponential or flocculent phase. Moreover, the decrease of Vmax of PFK or PK displayed the crucial role in down-regulation of flux in flocculent phase. In addition, the metabolic state of ale strain was more sensitive to the variation of temperature than that of lager strain. The results of the metabolic flux and nodes control analysis in brewer??s yeasts under different fermentation temperature may provide an alternative approach to regulate glycolytic flux by changing Vmax and improve the production efficiency and beer quality. 相似文献
Comb-like polymers with biocompatible oxyethylene backbones and amphiphilic side groups were synthesized via polymer-analogous reactions. Using these polymers, indomethacin-loaded polymeric micelles were fabricated with various drug-to-polymer weight ratios using the oil-in-water emulsion technique. In addition, the size, size distribution, CMC, drug-loading content, and entrapment efficiency of the polymeric micelles were analyzed. The volume-weighted diameters of polymeric micelles ranged from 10 to 140 nm and were narrowly distributed for passive targeting drug delivery. The CMCs were lower (approximately 10(-8) M) than for conventional surfactants and block copolymers. 相似文献