Interrupted adenylation domains are enigmatic fusions, in which one enzyme is inserted into another to form a highly unusual bifunctional enzyme. We present the first crystal structure of an interrupted adenylation domain that reveals a unique embedded methyltransferase. The structure and functional data provide insight into how these enzymes N-methylate amino acid precursors en route to nonribosomal peptides.

Stereodefined carbon-halogen bonds are ubiquitous in nature with several natural products exhibiting this motif.[1] While the biogenetic origins of this unique chiral functionality has been a subject of several investigations in the past, [2] attempts by organic chemists to forge the carbon-halogen bond stereoselectively have largely been unsuccessful. This problem has come into focus only recently. Several elegant reports of asymmetric halogenations of alkenes and alkynes followed by an intramolecular attack of a pendant nucleophile have appeared in the literature in the last decade. Kang et al. reported a cobalt-salen catalyzed iodoetherification reaction.[3a] An asymmetric fluorocyclization of allyl silanes mediated by a cinchona alkaloid dimer was reported by Gouverneur and co-workers.[3b] Tang and co-workers disclosed an asymmetric bromolactonization of enynes catalyzed by a cinchona alkaloid derived urea; other bromolactonizations have also appeared following the disclosure of their report.[3c-e] More recently, Veitch and Jacobsen reported an asymmetric iodolactonization reaction mediated by chiral thiourea catalysts.[3f] Polyene cyclizations induced by chiral halonium ions have also been realized as reported by the research groups of Ishihara and Snyder.[4a-c] However, given the fledgling nature of this research area, one may find it easy to highlight the many drawbacks and limitations even in the present state of the art-for example, the relatively large catalyst loadings (superstoichiometric quantities in some cases) to achieve meaningful levels of enantioselectivity or the lack of a robust catalytic system that can catalyze a number of diverse reactions rather than one specific reaction. Moreover, efficient asymmetric chlorocyclizations have remained underdeveloped. This situation is attributable, at least in part, to the highly reactive nature of chloronium ions, which are known to exist in equilibrium with the corresponding carbocation rather than exclusively as cyclic chloronium ions, [5a-c] thus making the development of chlorocyclizations a formidable challenge.Our research group has recently reported the catalytic asymmetric chlorolactonization of alkenoic acids.[7] Herein, we disclose the efficient halocyclization of unsaturated amides to furnish chiral heterocycles. Furthermore, these heterocycles have been transformed into useful chiral building blocks such as amino alcohols.Chiral heterocycles such as oxazolines and dihydrooxazines are commonly encountered motifs in natural products, [8a] molecules of pharmaceutical interest, [8b] and in several chiral ligands.[8c] Their syntheses, however, usually employ stoichiometric quantities of chiral amino alcohols. With only one precedented method to access these molecules in a catalytic asymmetric fashion, [6] we were intrigued by the possibility of one-step access to these versatile chiral heterocycles by a catalytic asymmetric halocyclization of easily accessed unsaturated amides.We chose the conversion of benzamide 1 into oxazoline 2 as our init...

Dimethylation of amino acids consists of an interesting and puzzling series of events that could be achieved, during nonribosomal peptide biosynthesis, either by a single adenylation (A) domain interrupted by a methyltransferase (M) domain or by the sequential action of two of such independent enzymes. Herein, to establish the method by which Nature N,S-dimethylates l-Cys, we studied its formation during thiochondrilline A biosynthesis by evaluating TioS(AMAT) and TioN(AMA). This study not only led to identification of the exact pathway followed in Nature by these two enzymes for N,S-dimethylation of l-Cys, but also revealed that a single interrupted A domain can N,N-dimethylate amino acids, a novel phenomenon in the nonribosomal peptide field. These findings offer important and useful insights for the development and engineering of novel interrupted A domain enzymes to serve, in the future, as tools for combinatorial biosynthesis.

Tuberculosis (TB) remains one of the leading causes of mortality worldwide. Hence, the identification of highly effective antitubercular drugs with novel modes of action is crucial. In this paper, we report the discovery and development of pyrrolo[1,5-a]pyrazine-based analogues as highly potent inhibitors of the Mycobacterium tuberculosis (Mtb) acetyltransferase enhanced intracellular survival (Eis), whose up-regulation causes clinically observed resistance to the aminoglycoside (AG) antibiotic kanamycin A (KAN). We performed a structure–activity relationship (SAR) study to optimize these compounds as potent Eis inhibitors both against purified enzyme and in mycobacterial cells. A crystal structure of Eis in complex with one of the most potent inhibitors reveals that the compound is bound to Eis in the AG binding pocket, serving as the structural basis for the SAR. These Eis inhibitors have no observed cytotoxicity to mammalian cells and are promising leads for the development of innovative AG adjuvant therapies against drug-resistant TB.

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A two-drug combination therapy where one drug targets an offending cell and the other targets a resistance mechanism to the first drug is a time-tested, yet underexploited approach to combat or prevent drug resistance. By high-throughput screening, we identified a sulfonamide scaffold that served as a pharmacophore to generate inhibitors of Mycobacterium tuberculosis acetyltransferase Eis, whose upregulation causes resistance to the aminoglycoside (AG) antibiotic kanamycin A (KAN) in Mycobacterium tuberculosis. Rational systematic derivatization of this scaffold to maximize Eis inhibition and abolish the Eis-mediated KAN resistance of M. tuberculosis yielded several highly potent agents. A crystal structure of Eis in complex with one of the most potent inhibitors revealed that the inhibitor bound Eis in the AG-binding pocket held by a conformationally malleable region of Eis (residues 28–37) bearing key hydrophobic residues. These Eis inhibitors are promising leads for preclinical development of innovative AG combination therapies against resistant TB.

A common cause of resistance to kanamycin (KAN) in tuberculosis is overexpression of the enhanced intracellular survival (Eis) protein. Eis is an acetyltransferase that multiacetylates KAN and other aminoglycosides, rendering them unable to bind the bacterial ribosome. By high-throughput screening, a series of substituted 1,2,4-triazino[5,6 b]indole-3-thioether molecules were identified as effective Eis inhibitors. Herein, we purchased 17 and synthesized 22 new compounds, evaluated their potency, and characterized their steady-state kinetics. Four inhibitors were found not only to inhibit Eis in vitro, but also to act as adjuvants of KAN and partially restore KAN sensitivity in a Mycobacterium tuberculosis KAN-resistant strain in which Eis is upregulated. A crystal structure of Eis in complex with a potent inhibitor and CoA shows that the inhibitors bind in the aminoglycoside binding site snugly inserted into a hydrophobic cavity. These inhibitors will undergo preclinical development as novel KAN adjuvant therapies to treat KAN-resistant tuberculosis.

In this article a new environmentally friendly catalytic method is described for the efficient monoiodination and bromination of arenes and also iodoetherification and iodolactonization of olefins using hydrogen peroxide as the terminal oxidant. The method is based on using sodium iodide or sodium bromide, hydrogen peroxide (35%) and ceriumA C H T U N G T R E N N U N G (III) chloride as an effective catalyst in water at room temperature or under reflux conditions. By this protocol, iodination of anilines proceeded with high regioselectivity at the para position with the formation of small amounts of the ortho isomers. However, bromination of anilines proceeded with absolute regioselectivity to give the para isomers as the sole products in high yields. Iodinations and brominations of m-xylene, toluene, chloro-and bromobenzenes were proceeded with excellent regioselectivity to produce the para isomers as the sole products. Benzene was also halogenated by this catalytic system to give the monohalogenated benzene in good yields. Iodoetherification and iodolactonization of olefins also proceeded easily in high yields at room temperature. However, the bromination of olefins by this protocol failed and the starting materials were detected intact.