A Switch for the transfer of substrate between nonribosomal peptide and polyketide modules of the rifamycin synthetase assembly line

A nonribosomal peptide synthetase (NRPS) loading module and a polyketide synthase (PKS) elongation module catalyze the preliminary steps in the biosynthesis of the rifamycin antibiotics. A benzoate molecule is covalently attached to the phosphopantetheine arm of the thiolation domain of the loading module when its reaction partner methylmalonyl-CoA is absent. Occupancy of the thiolation domain of the elongation module by a methylmalonyl moiety appears to trigger intermodular transfer of benzoate to the ketosynthase domain of the elongation module. This transthiolation event is fast relative to the initial loading of benzoate onto the loading module. It will be of interest to determine if these results are generally true for intermodular acyl transfer in other NRPS-PKS and PKS assembly lines.     

Proline-catalyzed asymmetric assembly reactions: enzyme-like assembly of carbohydrates and polyketides from three aldehyde substrates

Directed asymmetric assembly of simple achiral building blocks into stereochemically complex molecules like triketides has been described for the first time using l-proline catalyzed asymmetric double aldol reactions. The product pyranoses contain four asymmetric centers constructed under proline catalysis in a highly diastereoselective and modestly enantioselective fashion from three aldehyde molecules. These results suggest that the construction of complex products from simple starting materials is within the realm of organocatalysis involving the simple naturally occurring amino acid l-proline. Our successful assembly of pyranoses from simple aldehydes under proline catalysis suggests that this approach may warrant consideration as a prebiotic route to sugars and polyketides.     

Yersiniabactin synthetase: a four-protein assembly line producing the nonribosomal peptide/polyketide hybrid siderophore of Yersinia pestis

Yersiniabactin synthetase comprises four proteins, YbtE, HMWP1, HMWP2, and YbtU, encompassing seventeen functional domains, twelve catalytic and five carrier, to select, activate, and incorporate salicylate, three cysteines, and one malonyl moiety into the iron chelator yersiniabactin (Ybt). In the present study, yersiniabactin has been reconstituted in vitro from the 4 protein assembly line by the use of eight biosynthetic precursors. The rate of one turnover, comprising 22 chemical operations performed by the assembly line to release the completed Ybt molecule, was determined at 1.4 min(-1). During the course of Ybt production, the elongating acyl-S-enzyme chain was shown to transfer across a nonribosomal peptide synthetase/polyketide synthase (NRPS/PKS) interprotein interface and then a PKS/NRPS intraprotein interface. This study on the Ybt synthetase assembly line represents the first complete in vitro reconstitution of a nonribosomal peptide/polyketide hybrid system.     

Characterization of the cereulide NRPS alpha-hydroxy acid specifying modules: activation of alpha-keto acids and chiral reduction on the assembly line

Several nonribosomal peptide natural products are composites of alpha-hydroxy acid and alpha-amino acid monomers. Cereulide, the emetic toxin from the human pathogen Bacillus cereus, and valinomycin, from Streptomyces spp., are closely related macrocyclic K+ ionophores. The macrocyclic core of each natural product contains alternating peptide (six) and ester (six) bonds, and their cyclododecadepsipeptide structures consist of a tetradepsipeptide unit repeated three times. Here we overexpress the cereulide NRPS alpha-hydroxy acid specifying modules from CesA and CesB and demonstrate that each contains an alpha-keto acid activating adenylation domain and a chiral alpha-ketoacyl-S-carrier protein reductase (alpha-KR). The logic used by the cereulide NRPS is likely at work in the valinomycin NRPS and may be the general strategy used in bacterial NRPSs to form alpha-hydroxy acid containing natural products.     

Application of hitherto unexplored macrocyclization strategies in the epothilone series: novel epothilone analogs by total synthesis

A total synthesis of Epothilone 490 and a synthesis of 11-hydroxy dEpoB utilizing a vinyl-boronate cross-metathesis followed by a Suzuki macrocyclization. A mild route to reach aldehydes from terminal olefins, anticipating Nozaki-Kishi macrocyclization is described.     

Improved synthesis of epothilone B employing alkylation of an alkyne for assembly of subunits

[formula: see text] A strategy for assembling the two principal modules of epothilone B was developed that merges an allylic bromide with a terminal acetylene to fabricate the C10-C11 bond of the macrocycle. The resulting alkyne was semihydrogenated to give a seco ester previously employed in our total synthesis of epothilone B. This new approach affords a more efficient route to the naturally occurring macrolide and to its 9,10-dehydro analogue.     

Heterogeneous ultrathin films fabricated by alternate assembly of exfoliated layered double hydroxides and polyanion

Transparent heterogeneous ultrathin films of exfoliated layered double hydroxide (LDHs) nanosheets, fabricated alternately with polyanion, have been obtained via a layer-by-layer electrostatic self-assembly which yields a series of novel LDH films with potential multifunctionality.     

In vivo metabolism of epothilone B in tumor-bearing nude mice: identification of three new epothilone B metabolites by capillary high-pressure liquid chromatography/mass spectrometry/tandem mass spectrometry

Epothilone B is a 16-membered macrolide produced by the myxobacterium Sorangium cellulosum and is currently under clinical investigation. Experimentally, epothilone B demonstrates potent antiproliferative activity at the nanomolar level in vitro, and potent regression-producing antitumor activity in vivo, similar to paclitaxel (Taxol). In order to foster the design of improved derivatives, the potential biotransformation products of epothilone B formed in liver of tumor-bearing mice after intravenous administration of 10 mg/kg were characterized in an early stage of compound development. Solely on the basis of capillary high-performance liquid chromatography, combined either with electrospray tandem mass spectrometry (in precursor and product ion scan mode) and single analyzing time-of-flight mass spectrometry (H/D exchange and accurate mass measurement), three main metabolites could be detected. The three metabolites, formed by the liver, have in common that the epoxide ring was hydrolyzed and that the macrocyclic lactone ring was opened to the acid. In two cases it is assumed that open-chain intermediates re-cycled either to a lactone or, after conjugation with taurine, to the respective lactam. The proposed structures were additionally supported by the determination of the number of the exchangeable hydrogen atoms and by confirmation of the proposed elemental composition by exact mass measurement.     

Flame-retardant Coating by Alternate Assembly of Poly（vinylphosphonic acid） and Polyethylenimine for Ramie Fabrics

A novel intumescent flame retardant coating,consisting of poly(vinylphosphonic acid)(PVPA) as the acid source and branched polyethylenimine(BPEI) as the blowing agent,was constructed on the surface of ramie fabrics by alternate assembly to remarkably improve the flame retardancy of ramie.The PVPA/BPEI coating on the surface of individual fibers of ramie fabric pyrolyzes to form protective char layer upon heating/burning and improves the flame retardancy of ramie.Thermogravimetric analysis reveals that the PVPA/BPEI-coated ramie fabrics left as much as 25.8 wt% residue at 600 °C,while the control(uncoated) fabric left less than 1.4 wt% residue.Vertical flame test shows that all PVPA/BPEI-coated fabrics have shorter after-flame time,and the residues well preserved the original weave structure and fiber morphology,whereas,the uncoated fabric left only ashes.Microscale combustion calorimetry shows that the PVPA/BPEI coatings greatly reduce the total heat release by as much as 66% and the heat release capacity by 76%,relative to those of the uncoated fabric.     

Anion-directed assembly: the first fluoride-directed double helix

The crystal structures of anion complexes of two nitroaromatic functionalised isophthalamides are reported; the structures reveal assembly around anions in the solid-state and in the case of the fluoride complex of receptor 2, the formation of a double helix.     

Selective DNA-mediated assembly of gold nanoparticles on electroded substrates

Motivated by the technological possibilities of electronics and sensors based on gold nanoparticles (Au NPs), we investigate the selective assembly of such NPs on electrodes via DNA hybridization. Protocols are demonstrated for maximizing selectivity and coverage using 15mers as the active binding agents. Detailed studies of the dependences on time, ionic strength, and temperature are used to understand the underlying mechanisms and their limits. Under optimized conditions, coverage of Au NPs on Au electrodes patterned on silicon dioxide (SiO2) substrates was found to be approximately 25-35%. In all cases, Au NPs functionalized with non-complementary DNA show no attachment and essentially no nonspecific adsorption is observed by any Au NPs on the SiO2 surfaces of the patterned substrates. DNA-guided assembly of multilayers of NPs was also demonstrated and, as expected, found to further increase the coverage, with three deposition cycles resulting in a surface coverage of approximately 60%.     

Nanoparticle-mediated epitaxial assembly of colloidal crystals on patterned substrates

We have studied the assembly of 3-D colloidal crystals from binary mixtures of colloidal microspheres and highly charged nanoparticles on flat and epitaxially patterned substrates created by focused ion beam milling. The microspheres were settled onto these substrates from dilute binary mixtures. Laser scanning confocal microscopy was used to directly observe microsphere structural evolution during sedimentation, nanoparticle gelation, and subsequent drying. After microsphere settling, the nanoparticle solution surrounding the colloidal crystal was gelled in situ by introducing ammonia vapor, which increased the pH and enabled drying with minimal microsphere rearrangement. By infilling the dried colloidal crystals with an index-matched fluorescent dye solution, we generated full 3-D reconstructions of their structure including defects as a function of initial suspension composition and pitch of the patterned features. Through proper control over these important parameters, 3-D colloidal crystals were created with low defect densities suitable for use as templates for photonic crystals and photonic band gap materials.     

Total synthesis of epothilone a through stereospecific epoxidation of the p-methoxybenzyl ether of epothilone C

The total synthesis of epothilone A is described by the coupling four segments 4-7 a. Three of the segments, 4, 5 and 7 a, have only one chiral center; all other chiral centers were introduced by simple asymmetric catalytic reactions. The key steps are the ring opening of epoxide 5 with acetylide 8 for the construction of the C12-C13 cis double bond and a practical hydrolytic kinetic resolution (HKR) developed by Jacobsen group for the introduction the chiral center at C3. Especially, the stereospecific epoxidation of 3-O-PMB epothilone C 3 b through long-range effect of 3-O-PMB protecting group gave high yields of the C12-C13 alpha-epoxide for the synthesis of target molecule.