Journal: Journal of the American Chemical Society
Chemically modified proteins are invaluable tools for studying the molecular details of biological processes, and they also hold great potential as new therapeutic agents. Several methods have been developed for the site-specific modification of proteins, one of the most widely used being expressed protein ligation (EPL) in which a recombinant α-thioester is ligated to an N-terminal Cys-containing peptide. Despite the widespread use of EPL, the generation and isolation of the required recombinant protein α-thioesters remain challenging. We describe here a new method for the preparation and purification of recombinant protein α-thioesters using engineered versions of naturally split DnaE inteins. This family of autoprocessing enzymes is closely related to the inteins currently used for protein α-thioester generation, but they feature faster kinetics and are split into two inactive polypeptides that need to associate to become active. Taking advantage of the strong affinity between the two split intein fragments, we devised a streamlined procedure for the purification and generation of protein α-thioesters from cell lysates and applied this strategy for the semisynthesis of a variety of proteins including an acetylated histone and a site-specifically modified monoclonal antibody.
Selective catalytic synthesis of Z-olefins has been challenging. Here we describe a method to produce 1,2-disubstituted olefins in high Z selectivity via reductive cross-coupling of alkyl halides with terminal arylalkynes . The method employs inexpensive and non-toxic catalyst (iron(II) bromide) and reductant (zinc). The substrate scope encompasses primary, secondary, and tertiary alkyl halides, and the reaction tolerates a large number of functional groups. The utility of the method is demonstrated in the synthesis of several pharmaceutically relevant molecules. Mechanistic study suggests that the reaction proceeds through an iron-catalyzed anti-selective carbozincation pathway.
Complete structural elucidation of natural products is often challenging due to structural complexity and limited availability. This is true for present-day secondary metabolites, but even more for exceptionally preserved secondary metabolites of ancient organisms that potentially provide insights into the evolutionary history of natural products. Here we report the full structure and absolute configuration of the borolithochromes, enigmatic boron-containing pigments from a Jurassic putative red alga, from samples of less than 50 µg using microcryoprobe NMR, CD spectroscopy, and DFT calculations, and reveal their polyke-tide origin. The pigments are identified as spiroborates with two pentacyclic sec-butyl-trihydroxy-methyl-benzo[gh]tetraphen-one ligands and less substituted deriva-tives. The configuration of the sec-butyl group is found to be (S). Because the exceptional benzo[gh]tetraphene scaffold is otherwise only observed in the recently discovered polyke-tide clostrubin from a present-day Clostridium bacterium, the Jurassic borolithochromes now can be unambiguously linked to the modern polyketide, providing evidence that the fossil pigments are almost originally preserved secondary metabolites and suggesting that the pigments in fact may have been produced by an ancient bacterium. The borolithochromes differ fundamentally from previously described boronated polyketides and represent the first boronated aromatic polyketides found so far. Our results demonstrate the potential of microcryoprobe NMR in the analysis of previously little-explored secondary metabolites from ancient organisms and reveal the evolutionary significance of clostrubin-type polyketides.
Superomniphobic surfaces display contact angles > 150° and low contact angle hysteresis with essentially all contacting liquids. In this work, we report surfaces that display superomniphobicity with a range of different non-Newtonian liquids, in addition to super-omniphobicity with a wide range of Newtonian liquids. Our surfaces possess hierarchical scales of re-entrant texture that significantly reduces the solid-liquid contact area. Virtually all liquids including concentrated organic and inorganic acids, bases and solvents, as well as, vis-coelastic polymer solutions can easily roll-off and bounce on our surfaces. Consequently, they serve as effective chemical shields against virtually all liquids - organic or inorganic, polar or non-polar, Newtonian or non-Newtonian.
Lithium-ion conducting solid electrolytes hold the promise for enabling high-energy battery chemistries and circumventing safety issues of conventional lithium batteries. Achieving the combination of high ionic conductivity and broad electrochemical window in solid electrolytes is a grand challenge for the synthesis of battery materials. Herein we show an enhancement of room-temperature lithium-ion conductivity of 3 orders of magnitude by creating nanostructured Li3PS4. This material has a wide (5 V) electrochemical window and superior chemical stability against lithium metal. The nanoporous structure of Li3PS4 reconciles two vital effects that enhance ionic conductivity: (1) The reduced dimension to nanometer-sized framework stabilizes the high conduction beta phase that occurs at elevated temperatures; and (2) The high surface-to-bulk ratio of nanoporous β-Li3PS4 promotes surface conduction. Manipulating the ionic conductivity of solid electrolytes has far-reaching implications for materials design and synthesis in a broad range of applications such as batteries, fuel-cells, sensors, photovoltaic systems, and so forth.
We report selective electrocatalytic reduction of carbon dioxide (CO2) to carbon monoxide (CO) on gold (Au) nanoparticles (NPs) in 0.5 M KHCO3 at 25℃. Among monodisperse 4-, 6-, 8-, and 10-nm NPs tested, the 8 nm Au NPs show the maximum Faradaic efficiency (FE) (up to 90% at -0.67 V vs. reversible hydrogen electrode, RHE). Density functional theory (DFT) calculations suggest that the presence of dominant edge sites over corner sites (active for the competitive H2 evolution reaction) on the Au NP surface facilitates the stabilization of the reduction intermediates, such as COOH*, and the formation of CO. This mechanism is further supported by the fact that Au NPs embedded in a matrix of butyl-3-methylimidazolium hexafluorophosphate for more efficient COOH* stabilization exhibit even higher reaction activity (3 A/g mass activity) and selectivity (97% FE) at -0.52 V (vs. RHE). The work demonstrates the great potentials of using monodisperse Au NPs to optimize the available reaction intermediate binding sites for efficient and selective electrocatalytic reduction of CO2 to CO.
Iron sulfide generated in situ from elemental sulfur and iron was found to be highly efficient to catalyze redox/condensation cascade reaction between 2-amino/hydroxy nitrobenzenes and activated methyl groups. This method represents a straightforward and highly atom economical approach to 2-hetaryl- benzimidazoles and benzoxazoles.
Heroin is a highly abused opioid and incurs a significant detriment to society worldwide. In an effort to expand the limited pharmacotherapy options for opioid use disorders, a heroin conjugate vaccine was developed through comprehensive evaluation of hapten structure, carrier protein, adjuvant and dosing. Immunization of mice with an optimized heroin-tetanus toxoid (TT) conjugate formulated with adjuvants alum and CpG oligodeoxynucleotide (ODN) generated heroin ‘immunoantagonism’, reducing heroin potency by >15-fold. Moreover, the vaccine effects proved to be durable, persisting for over eight months. The lead vaccine was effective in rhesus monkeys, generating significant and sustained anti-drug IgG titers in each subject. Characterization of both mouse and monkey anti-heroin antibodies by surface plasmon resonance (SPR) revealed low nanomolar antiserum affinity for the key heroin metabolite, 6-acetylmorphine (6AM), with minimal cross reactivity to clinically-used opioids. Following a series of heroin challenges over six months in vaccinated monkeys, drug-sequestering antibodies caused marked attenuation of heroin potency (>4-fold) in a schedule-controlled responding (SCR) behavioral assay. Overall, these preclinical results provide an empirical foundation supporting the further evaluation and potential clinical utility of an effective heroin vaccine in treating opioid use disorders.
The ability to detect DNA modification sites at single base resolution could significantly advance studies regarding DNA adduct levels, which are extremely difficult to determine. Artificial nucleotides that are specifically incorporated opposite a modified DNA site offer a potential strategy for detection of such sites by DNA polymerase-based systems. Here we investigated newly synthesized base-modified benzimidazole-derived 2´-deoxynucleoside-5´-O-triphosphates in their action on DNA polymerases when performing translesion DNA synthesis past the pro-mutagenic DNA adduct O(6)-benzylguanine (O(6)-BnG). We found that a mutated form of KlenTaq DNA polymerase, i.e. KTqM747K catalyzed O(6)-BnG adduct-specific processing of the artificial BenziTP in favor of the natural dNTPs. Steady-state kinetic parameters revealed that KTqM747K catalysis of BenziTP is 25-fold more efficient for template O(6)-BnG than G, and 5-fold more efficient than natural dTMP misincorporation in adduct bypass. Furthermore, the nucleotide analog BenziTP is required for full-length product formation in O(6)-BnG bypass, as without BenziTP the polymerase stalls at the adduct site. By combining the KTqM747K polymerase and BenziTP, a first round of DNA synthesis enabled subsequent amplification of Benzi-containing DNA. These results advance the development of technologies for detecting DNA adducts.
The total synthesis of the Lycopodium alkaloid lyconadin A was accomplished and it was applied to the total syntheses of the related congeners, lyconadins B, and C. Lyconadin A has attracted attention as a challenging target for total synthesis due to the unprecedented pentacyclic skeleton. Our synthesis of lyconadin A features a facile construction of the highly fused tetracyclic skeleton through a combination of an aza-Prins reaction and an electrocyclic ring opening followed by a formation of a C-N bond. Transformation of the bromoalkene moiety of the tetracycle to a key enone intermediate was extensively investigated, and three methods via sulfide, oxime, or azide intermediates were established. A pyridone ring was constructed from the key enone interme-diate to complete the synthesis of lyconadin A. A dihydropyridone ring could also be formed from the same enone intermediate, leading to a synthesis of lyconadin B. Establishment of the conditions for an electrocyclic ring opening without formation of the C-N bond resulted in completion of the total synthesis of lyconadin C.