Fluorescent labeling of RNA and DNA on the Hoogsteen edge using sulfinate chemistry.

We have devised a single pot, low-cost method to add azide groups to unmodified nucleic acids without the need for enzymes or chemically modified nucleoside triphosphates. This involves reacting an azide-containing sulfinate salt with the nucleic acid, leading to replacement of C-H bonds on the nucleobase aromatic rings with C-R, where R is the azide-containing linker derived from the original sulfinate salt. With the addition of azide functional groups, the modified nucleic acid can easily be reacted with any alkyne-labeled compound of interest, including fluorescent dyes as shown in this work. This methodology enables the fluorescent labeling of a wide variety of nucleic acids, including natively folded RNAs, under mild conditions with minimal effects upon biochemical function and ribozyme catalysis. To demonstrate this, we show that a pair of labeled complementary ssDNA oligonucleotides (oligos) can hybridize to form dsDNA, even when labeled with multiple fluorophores per oligo. In addition, we also demonstrate that two different group II introns can splice when prelabeled internally with fluorophores, using our method. Broadly, this demonstrates that sulfinate modification of RNA is compatible with ribozyme function and Watson-Crick pairing, while preserving the labile backbone.

Identification of a novel enzyme and the regulation of key enzymes in mammalian taurine synthesis.

Taurine has many pharmacological roles on various tissues. The maintenance of abundant taurine content in the mammalian body through endogenous synthesis, in addition to exogenous intake, is the essential factor for morphological and functional maintenances in most tissues. The synthesis of taurine from sulfur-containing amino acids is influenced by various factors. Previous literature findings indicate the influence of the intake of proteins and sulfur-containing amino acids on the activity of the rate-limiting enzymes cysteine dioxygenase and cysteine sulfinate decarboxylase. In addition, the regulation of the activity and expression of taurine-synthesis enzymes by hormones, bile acids, and inflammatory cytokines through nuclear receptors have been reported in liver and reproductive tissues. Furthermore, flavin-containing monooxygenase subtype 1 was recently identified as the taurine-synthesis enzyme that converts hypotaurine to taurine. This review introduces the novel taurine synthesis enzyme and the nuclear receptor-associated regulation of key enzymes in taurine synthesis.

Carbenoids with Sulfinate as Nucleofuge for Matteson-Type Homologation: Direct Insertion of Oxygen- and Nitrogen-Substituted Units into Carbon‒Boron Bonds.

Carbenoid insertion into boronate carbon-boron bonds, namely the Matteson-type homologation, has been recognized as a powerful tool for constructing carbon-carbon bonds. However, some limitations and inconvenience still exist with the carbenoids currently employed, such as the use of highly cryogenic and basic conditions. Herein, we report a new class of stable carbenoids with sulfinate as nucleofuge for Matteson-type homologations, which directly introduce O- and N-substituted methylenes into carbon-boron bonds. Enabled by oxazaborolidines as the boronic substrates, the reaction is operatable at 0 oC or room temperature with weaker bases. Broad functional groups, including acidic C-H bonds, can be tolerated. The synthetic utility of this method has been demonstrated in the gram-scale synthesis and iterative insertion of various carbenoids.

Testosterone enhances taurine synthesis by upregulating androgen receptor and cysteine sulfinic acid decarboxylase expressions in male mouse liver.

Taurine is an end-product of cysteine metabolism, whereas cysteine dioxygenase (CDO) and cysteine sulfinate decarboxylase (CSAD) are key enzymes regulating taurine synthesis. Sex steroids, including estrogens and androgens, are associated with liver physiopathological processes; however, we still do not know whether taurine and sex steroids interact in regulating liver physiology and hepatic diseases, and whether there are sex differences, although our recent study shows that the estrogen is involved in regulating taurine synthesis in mouse liver. The present study was thus proposed to identify whether 17-ß-estradiol and testosterone (T) play their roles by regulating CDO and CSAD expression and taurine synthesis in male mouse liver. Our results demonstrated that testosterone did not have a significant influence on CDO expression but significantly enhanced CSAD, androgen receptor (AR) expressions, and taurine levels in mouse liver, cultured hepatocytes, and HepG2 cells, whereas these effects were abrogated by AR antagonist flutamide. Furthermore, our results showed that testosterone increased CSAD-promoter-luciferase activity through the direct interaction of the AR DNA binding domain with the CSAD promoter. These findings first demonstrate that testosterone acts as an important factor to regulate sulfur amino acid metabolism and taurine synthesis through AR/CSAD signaling pathway. In addition, the in vivo and in vitro experiments showed that 17-ß-estradiol has no significant effects on liver CSAD expression and taurine synthesis in male mice and suggest that the effects of sex steroids on the taurine synthesis in mouse liver have sex differences. These results are crucial for understanding the physiological functions of taurine/androgen and their interacting mechanisms in the liver.NEW & NOTEWORTHY This study demonstrates that testosterone functions to enhance taurine synthesis by interacting with androgen receptor and binding to cysteine sulfinate decarboxylase (CSAD) promoter zone. Whereas estrogen has no significant effects either on liver CSAD expression or taurine synthesis in male mice and suggests that the effects of sex steroids on taurine synthesis in the liver have gender differences. These new findings are the potential for establishing effective protective and therapeutic strategies for liver diseases.

Synthesis and Antimalarial Evaluation of Halogenated Analogues of Thiaplakortone A.

The incorporation of bromine, iodine or fluorine into the tricyclic core structure of thiaplakortone A (1), a potent antimalarial marine natural product, is reported. Although yields were low, it was possible to synthesise a small nine-membered library using the previously synthesised Boc-protected thiaplakortone A (2) as a scaffold for late-stage functionalisation. The new thiaplakortone A analogues (3-11) were generated using N-bromosuccinimide, N-iodosuccinimide or a Diversinate™ reagent. The chemical structures of all new analogues were fully characterised by 1D/2D NMR, UV, IR and MS data analyses. All compounds were evaluated for their antimalarial activity against Plasmodium falciparum 3D7 (drug-sensitive) and Dd2 (drug-resistant) strains. Incorporation of halogens at positions 2 and 7 of the thiaplakortone A scaffold was shown to reduce antimalarial activity compared to the natural product. Of the new compounds, the mono-brominated analogue (compound 5) displayed the best antimalarial activity with IC50 values of 0.559 and 0.058 µM against P. falciparum 3D7 and Dd2, respectively, with minimal toxicity against a human cell line (HEK293) observed at 80 µM. Of note, the majority of the halogenated compounds showed greater efficacy against the P. falciparum drug-resistant strain.

Negative allosteric modulators of NMDA receptors with GluN2B subunit: Alanine-derived benzoxazolone bioisosteres of 2-methyl-3-benzazepine-1,7-diols.

Inspired by besonprodil, the phenol of potent negative allosteric modulators of GluN2B-N-methyl-d-aspartate (NMDA) receptors was replaced by a benzoxazolone system. To increase the similarity to the lead compounds, an additional methyl moiety was installed in the 8-position of tricyclic oxazolobenzazepines, resulting in compounds 6. The additional methyl moiety originates from alanine, which was introduced by a Mitsunobu reaction of benzoxazolylethanol 7 with N-triflyl-protected alanine methyl ester. A crucial feature of the synthesis was the protection of the oxazolone ring by an allyl moiety, which was cleaved off at the end of the synthesis by RhCl3 -catalyzed isomerization. Due to the additional methyl moiety, the intramolecular Friedel-Crafts acylation of acid 10 to afford ketone 11 required careful optimization to minimize the formation of the side product tetrahydroisoquinoline 16. Alkylation or reductive alkylation of secondary amine 13 led to diastereomeric oxazolobenzazepines cis-14 and trans-14, which were separated by flash chromatography. Phenylbutyl derivatives cis-6a and trans-6a revealed twofold higher GluN2B affinity than analog 5a without 8-CH3 group. The methylated oxazolobenzazepines 6 and 14 did not interact with the phencyclidine binding site of NMDA receptors and σ2 receptors. However, the σ1 receptor preferred cis-configured oxazolobenzazepines. The highest σ1 receptor affinities were obtained for cis-14a (Ki = 26 nM) and cis-6b (Ki = 30 nM).

Reductant-Free Cross-Electrophile Synthesis of Di(hetero)arylmethanes by Palladium-Catalyzed Desulfinative C-C Coupling.

An efficient Pd-catalyzed one-pot desulfinative cross-coupling to access medicinally relevant di(hetero)arylmethanes is reported. The method is reductant-free, and involves a sulfinate transfer reagent and a Pd-catalyst mediating the union of two electrophilic coupling partners; a (hetero)aryl halide and a benzyl halide. We establish for the first time that benzyl sulfinates, generated in situ, undergo efficient Pd-catalyzed desulfinative cross-coupling with (hetero)aryl halides to generate di(hetero)arylmethanes. The reaction can be extended to benzylic pseudohalides derived from benzyl alcohols. The reactions are straightforward to perform and scalable, and all reaction components are commercially available.

1,1-Diphenylvinylsulfide as a Functional AIEgen Derived from the Aggregation-Caused-Quenching Molecule 1,1-Diphenylethene through Simple Thioetherification.

An efficient and readily scalable thioetherification between 1,1-diphenylethene (DPE) and sodium arylsulfinate was developed for the synthesis of 1,1-diphenylvinylsulfide (DPVS) with the yield up to 99 %. The photophysical properties of DPVS show that the introduction of arylsulfenyl groups onto the parent molecule DPE makes DPVS a novel type of aggregation-induced emission (AIE) luminogen (AIEgen) with large Stoke's shift (up to 188 nm). These DPVS possess AIE properties due to restriction of intramolecular motions (RIM), as demonstrated by crystal structure analysis. Importantly, the AIE performance of DPVS can be applied to sense the nitroaromatic explosive picric acid in aqueous systems through a "turn-off" response.

Reaction of Papaverine with Baran DiversinatesTM.

The reaction of papaverine with a series of Baran DiversinatesTM is reported. Although the yields were low, it was possible to synthesize a small biodiscovery library using this plant alkaloid as a scaffold for late-stage C-H functionalization. Ten papaverine analogues (2-11), including seven new compounds, were synthesized. An unexpected radical-induced exchange reaction is reported where the dimethoxybenzyl group of papaverine was replaced by an alkyl group. This side reaction enabled the synthesis of additional novel fragments based on the isoquinoline scaffold, which is present in numerous natural products. Possible reasons for the poor yields in the DiversinateTM reactions with this particular scaffold are discussed.

Molecular Characterization and Directed Evolution of a Metagenome-Derived l-Cysteine Sulfinate Decarboxylase.

l-Cysteine sulfinate decarboxylase (CSD, EC 4.1.1.29), the rate-limiting enzyme in taurine synthesis pathway, catalyzes l-cysteine sulfinic acid to form hypotaurine. Identification of the novel CSD that could improve the biosynthetic efficiency of taurine is important. An unexplored decarboxylase gene named undec1A was identified in a previous work through sequence-based screening of uncultured soil microorganisms. Random mutagenesis through sequential error-prone polymerase chain reaction was used in Undec1A. A mutant Undec1A-1180, which was obtained from mutagenesis library, had 5.62-fold higher specific activity than Undec1A at 35 °C and pH=7.0. Molecular docking results indicated that amino acid residues Ala235, Val237, Asp239, Ile267, Ala268, and Lys298 in the Undec1A-1180 protein helped recognize and catalyze the substrate molecules of l-cysteine sulfinic acid. These results could serve as a basis for elucidating the characteristics of the Undec1A-1180. Directed evolution technology is a convenient way to improve the biotechnological applications of metagenome-derived genes.