Plants grown at high densities perceive a decrease in the red to far-red (R:FR) ratio of incoming light, resulting from absorption of red light by canopy leaves and reflection of far-red light from neighboring plants. These changes in light quality trigger a series of responses known collectively as the shade avoidance syndrome. During shade avoidance, stems elongate at the expense of leaf and storage organ expansion, branching is inhibited, and flowering is accelerated. We identified several loci in Arabidopsis, mutations in which lead to plants defective in multiple shade avoidance responses. Here we describe TAA1, an aminotransferase, and show that TAA1 catalyzes the formation of indole-3-pyruvic acid (IPA) from L-tryptophan (L-Trp), the first step in a previously proposed, but uncharacterized, auxin biosynthetic pathway. This pathway is rapidly deployed to synthesize auxin at the high levels required to initiate the multiple changes in body plan associated with shade avoidance.
The benzothiazinone lead compound, BTZ043, kills Mycobacterium tuberculosis by inhibiting the essential flavo-enzyme DprE1, decaprenylphosphoryl-beta-D-ribose 2-epimerase. Here, we synthesized a new series of piperazine-containing benzothiazinones (PBTZ) and show that, like BTZ043, the preclinical candidate PBTZ169 binds covalently to DprE1. The crystal structure of the DprE1-PBTZ169 complex reveals formation of a semimercaptal adduct with Cys387 in the active site and explains the irreversible inactivation of the enzyme. Compared to BTZ043, PBTZ169 has improved potency, safety and efficacy in zebrafish and mouse models of tuberculosis (TB). When combined with other TB drugs, PBTZ169 showed additive activity against M. tuberculosis in vitro except with bedaquiline (BDQ) where synergy was observed. A new regimen comprising PBTZ169, BDQ and pyrazinamide was found to be more efficacious than the standard three drug treatment in a murine model of chronic disease. PBTZ169 is thus an attractive drug candidate to treat TB in humans.Subject Categories Microbiology, Virology & Host Pathogen Interaction; Pharmacology & Drug Discovery
Halogen atom incorporation into a scaffold of bioactive compounds often amplifies biological activity, as is the case for the anticancer agent salinosporamide A (1), a chlorinated natural product from the marine bacterium Salinispora tropica. Significant effort in understanding enzymatic chlorination shows that oxidative routes predominate to form reactive electrophilic or radical chlorine species. Here we report the genetic, biochemical and structural characterization of the chlorinase SalL, which halogenates S-adenosyl-L-methionine (2) with chloride to generate 5′-chloro-5′-deoxyadenosine (3) and L-methionine (4) in a rarely observed nucleophilic substitution strategy analogous to that of Streptomyces cattleya fluorinase. Further metabolic tailoring produces a halogenated polyketide synthase substrate specific for salinosporamide A biosynthesis. SalL also accepts bromide and iodide as substrates, but not fluoride. High-resolution crystal structures of SalL and active site mutants complexed with substrates and products support the S N 2 nucleophilic substitution mechanism and further illuminate halide specificity in this newly discovered halogenase family.The chlorinated marine natural product salinosporamide A (SalA, 1), a potent 20S proteasome inhibitor currently in phase 1 human clinical trials for the treatment of multiple myeloma and other cancers, is 500 times more active than its deschloro analog salinosporamide B (SalB, 5) ( Fig. 1) [1][2][3][4][5] . The biosynthetic origin of SalA's pharmacologically active chloroethyl side chain from an unknown sugar precursor differs from the analogous ethyl group in SalB, which instead © 2007 Nature Publishing Group Correspondence should be addressed to B.S.M. (bsmoore@ucsd.edu). 4 These authors contributed equally to this work. AUTHOR CONTRIBUTIONS A.S.E. and F.P. contributed equally to this paper. A.S.E. performed the genetic and biochemical experiments, F.P. and A.S.E. crystallized SalL, F.P. determined the structure and performed the sedimentation velocity studies. All authors discussed the results and wrote and commented on the manuscript.Reprints and permissions information is available online at http://npg.nature.com/reprintsandpermissions Accession codes. Protein Data Bank: The atomic coordinates and structure factors were deposited under PDB accession codes 2Q6I, 2Q6K, 2Q6O and 2Q6L for SalL wild-type complex with 5′-ClDA and L-methionine, SalL wild-type complex with adenosine, SalL Y70T complex with chloride and SAM, and SalL Y70T G131S complex with 5′-ClDA and L-methionine, respectively. S. cattleya fluorinase structures were deposited as part of previous studies under PDB codes 1RQP and 2C2W.Note: Supplementary information and chemical compound information is available on the Nature Chemical Biology website. HHMI Author Manuscript HHMI Author Manuscript HHMI Author Manuscriptoriginates from butyrate (6) 6 . Analysis of the 41-kilobase sal biosynthetic gene cluster from S. tropica revealed a polyketide synthase-nonribosomal peptide synthetase (PK...
Specialized metabolic enzymes biosynthesize chemicals of ecological importance, often sharing a pedigree with primary metabolic enzymes1. However, the lineage of the enzyme chalcone isomerase (CHI) remained a quandary. In vascular plants, CHI-catalyzed conversion of chalcones to chiral (S)-flavanones is a committed step in the production of plant flavonoids, compounds that contribute to attraction, defense2, and development3. CHI operates near the diffusion limit with stereospecific control4,5. While associated primarily with plants, the CHI-fold occurs in several other eukaryotic lineages and in some bacteria. Here we report crystal structures, ligand-binding properties, and in vivo functional characterization of a non-catalytic CHI-fold family from plants. A. thaliana contains five actively transcribed CHI-fold genes, three of which additionally encode amino-terminal chloroplast-transit sequences (cTP). These three CHI-fold proteins localize to plastids, the site of de novo fatty acid (FA) biosynthesis in plant cells. Furthermore, their expression profiles correlate with those of core FA biosynthetic enzymes, with maximal expression occurring in seeds and coinciding with increased FA storage in the developing embryo. In vitro, these proteins are Fatty Acid-binding Proteins (FAP). FAP knockout A. thaliana plants exhibit elevated alpha-linolenic acid levels and marked reproductive defects, including aberrant seed formation. Notably, the FAP discovery defines the adaptive evolution of a stereospecific and catalytically ‘perfected’ enzyme6 from a non-enzymatic ancestor over a defined period of plant evolution.
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