High throughput photo-oxidations in a packed bed reactor system.

The efficiency gains produced by continuous-flow systems in conducting photochemical transformations have been extensively demonstrated. Recently, these systems have been used in developing safe and efficient methods for photo-oxidations using singlet oxygen generated by photosensitizers. Much of the previous work has focused on the use of homogeneous photocatalysts. The development of a unique, packed-bed photoreactor system using immobilized rose bengal expands these capabilities as this robust photocatalyst allows access to and elaboration from these highly useful building blocks without the need for further purification. With this platform we were able to demonstrate a wide scope of singlet oxygen ene, [4+2] cycloadditions and heteroatom oxidations. Furthermore, we applied this method as a strategic element in the synthesis of the high-volume antimalarial artemisinin.

A convergent approach to the total synthesis of telmisartan via a Suzuki cross-coupling reaction between two functionalized benzimidazoles.

A direct and efficient total synthesis has been developed for telmisartan, a widely prescribed treatment for hypertension. This approach brings together two functionalized benzimidazoles using a high-yielding Suzuki reaction that can be catalyzed by either a homogeneous palladium source or graphene-supported palladium nanoparticles. The ability to perform the cross-coupling reaction was facilitated by the regio-controlled preparation of the 2-bromo-1-methylbenzimidazole precursor. This convergent approach provides telmisartan in an overall yield of 72% while circumventing many issues associated with previously reported processes.

Biochemical determination of enzyme-bound metabolites: preferential accumulation of a programmed octaketide on the enediyne polyketide synthase CalE8.

Despite considerable interest in the enediyne family of antitumor antibiotics, assembly of their polyketide core structures in nature remains poorly understood. Discriminating methods to access enzyme-bound intermediates are critical for elucidating unresolved polyketide and nonribosomal peptide biosynthetic pathways. Here, we describe the development of broadly applicable techniques for the mild chemical release and analysis of intermediates bound to carrier proteins (CPs), providing access to these species even in sensitive systems. These techniques were applied to CalE8, the polyketide synthase (PKS) involved in calicheamicin biosynthesis, facilitating the unambiguous identification of enzyme-bound polyketides on an enediyne PKS. Moreover, these methods enabled the preparation of fully unloaded CalE8, providing a "clean slate" for reconstituted activity and allowing us to demonstrate the preferential accumulation of a PKS-bound octaketide with evidence of programmed processing control by CalE8. This intermediate, which has the expected chain length for enediyne core construction, could previously only be indirectly inferred. These studies prove that this polyketide is an authentic product of CalE8 and may be a key precursor to the enediyne core of calicheamicin, as it is the only programmed, enzyme-bound species observed for any enediyne system to date. Our experimental advances into a generally inaccessible system illustrate the utility of these techniques for investigating CP-based biosynthetic pathways.

Interrogation of global active site occupancy of a fungal iterative polyketide synthase reveals strategies for maintaining biosynthetic fidelity.

Nonreducing iterative polyketide synthases (NR-PKSs) are responsible for assembling the core of fungal aromatic natural products with diverse biological properties. Despite recent advances in the field, many mechanistic details of polyketide assembly by these megasynthases remain unknown. To expand our understanding of substrate loading, polyketide elongation, cyclization, and product release, active site occupancy and product output were explored by Fourier transform mass spectrometry using the norsolorinic acid anthrone-producing polyketide synthase, PksA, from the aflatoxin biosynthetic pathway in Aspergillus parasiticus. Here we report the simultaneous observation of covalent intermediates from all catalytic domains of PksA from in vitro reconstitution reactions. The data provide snapshots of iterative catalysis and reveal an underappreciated editing function for the C-terminal thioesterase domain beyond its recently established synthetic role in Claisen/Dieckmann cyclization and product release. The specificity of thioesterase catalyzed hydrolysis was explored using biosynthetically relevant protein-bound and small molecule acyl substrates and demonstrated activity against hexanoyl and acetyl, but not malonyl. Processivity of polyketide extension was supported by the inability of a nonhydrolyzable malonyl analog to trap products of intermediate chain lengths and by the detection of only fully extended species observed covalently bound to, and as the predominant products released by, PksA. High occupancy of the malonyl transacylase domain and fast relative rate of malonyl transfer compared to starter unit transfer indicate that rapid loading of extension units onto the carrier domain facilitates efficient chain extension in a manner kinetically favorable to ultimate product formation.

Environmental control of the calicheamicin polyketide synthase leads to detection of a programmed octaketide and a proposal for enediyne biosynthesis.

A light in the dark: the fermentation products of the polyketide synthase CalE8 (without its cognate thioesterase) were identified and gave some insight into the elusive early steps of calicheamicin biosynthesis. Fermentation in either the light or dark resulted in different proportions of a new octaketide and led to an updated mechanistic proposal.

Production of octaketide polyenes by the calicheamicin polyketide synthase CalE8: implications for the biosynthesis of enediyne core structures.

Enediyne antibiotics are categorized according to the presence of either a 9- or 10-membered ring within their polyketide-derived core structures. Recent literature reports have favored the notion that biosynthetic divergence of the two structural families is determined by the enediyne polyketide synthases (PKSs) alone. We now disclose the simultaneous in vitro production of three octaketide polyenes by biosynthetic enzymes for the 10-membered enediyne calicheamicin gamma(1)(I), including the elusive beta-keto acid precursor to a previously described C15 methyl hexaenone. Alongside these two polyene products, we have additionally detected a hydrocarbon heptaene previously isolated only from 9-membered enediyne systems. The discovery of the heptaene in the calicheamicin system promotes a more convergent model for the early steps of enediyne biosynthesis. Furthermore, the synthesis of this set of octaketides by the enediyne PKS CalE8 and thioesterase CalE7 suggests, in contrast to recent biosynthetic proposals, that accessory enzymes may be necessary to initiate differentiation to 9- or 10-membered enediyne precursors, either by modulation of enediyne PKS activity or by interception and modification of polyketide chain-extension intermediates.

Analysis of the cercosporin polyketide synthase CTB1 reveals a new fungal thioesterase function.

The polyketide synthase CTB1 is demonstrated to catalyze pyrone formation thereby expanding the known biosynthetic repertoire of thioesterase domains in iterative, non-reducing polyketide synthases.

Simple, rapid procedure for the synthesis of chloromethyl methyl ether and other chloro alkyl ethers.

[Reaction: see text]. Zinc(II) salts catalyze the reaction between acetals and acid halides to provide haloalkyl ethers in near-quantitative yield. Reactions from millimole to mole scale are typically complete in 1-4 h with 0.01 mol % catalyst. The solutions of haloalkyl ethers thus obtained can be utilized directly in reactions in which the presence of the ester byproduct does not interfere. Excess haloalkyl ether is destroyed on workup, thereby minimizing exposure to this class of carcinogenic compounds.