The crystal structure of Rv0793, a hypothetical monooxygenase from M.␣tuberculosis

Lemieux, M. Joanne; Ference, C.; Cherney, M.M.; Wang, Metian; Garen, Craig R.; James, Michael N.G.

doi:10.1007/s10969-005-9004-6

Cited by 15 publications

(11 citation statements)

References 34 publications

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“…The transcript of several genes belonging to either PE/PPE or toxin-antitoxin modules or lipid metabolism were also downregulated in the mutant strains. We also observed that the transcript levels of Rv0793 (gene neighbouring to Rv0792c) and mymA operon (shown to be upregulated in acidic conditions) were increased in the mutant strain (50,67). These findings clearly suggest that Rv0792c regulates the expression of "subset" of genes that enables the bacteria to adapt and persist in host tissues.…”

Section: Discussionsupporting

confidence: 53%

Structural and functional characterization of Rv0792c from Mycobacterium tuberculosis: identifying small molecule inhibitors against GntR protein

Chauhan

Anand

Sharma

et al. 2021

Preprint

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In order to adapt in host tissues, microbial pathogens regulate their gene expression through an array of transcription factors. Here, we have functionally characterized Rv0792c, a GntR homolog from M. tuberculosis. In comparison to the parental strain, ΔRv0792c mutant strain of M. tuberculosis was compromised for survival upon exposure to oxidative stress, cell wall agents and infection in guinea pigs. RNA-seq analysis revealed that Rv0792c regulates the expression of genes that are involved in stress adaptation and virulence of M. tuberculosis. Solution small angle X-ray scattering (SAXS) data steered model building confirmed that the C-terminal region plays a pivotal role in dimer formation. Systematic evolution of ligands by exponential enrichment resulted in identification of ssDNA aptamers that can be used as a tool to identify small molecule inhibitors targeting Rv0792c. Using SELEX and SAXS data based modelling, we identified residues essential for the DNA binding activity of Rv0792c and I-OMe-Tyrphostin as an inhibitor of Rv0792c aptamer binding activity. Taken together, we provide a detailed shape-function characterization of GntR family of transcription factors from M. tuberculosis. To the best of our knowledge, this is the first study that has resulted in the identification of small molecule inhibitors against GntR family of transcription factors from bacterial pathogens.

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Section: Discussionsupporting

confidence: 53%

Structural and functional characterization of Rv0792c from Mycobacterium tuberculosis: identifying small molecule inhibitors against GntR protein

Chauhan

Anand

Sharma

et al. 2021

Preprint

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“…The three‐dimensional structure of OAC adopts almost the same overall structure observed in the plant and bacterial DABB proteins. A structure‐based similarity search revealed that the overall structure of the OAC–OA binary complex exhibits rmsd values of 1.1 Å, 1.4 Å and 2.5 Å with the plant DABB proteins AtHS1 (PDB code http://www.rcsb.org/pdb/search/structidSearch.do?structureId=1Q4R, 48% identity with OAC) , SP1 (PDB code http://www.rcsb.org/pdb/search/structidSearch.do?structureId=1TR0, 38% identity with OAC) and At5g22580 (PDB code http://www.rcsb.org/pdb/search/structidSearch.do?structureId=1RJJ, 32% identity with OAC) , and of 1.8 Å, 2.2 Å, 2.3 Å, 2.4 Å, 2.4 Å and 2.9 Å with the bacterial DABB proteins S. nogalater SnoaB (PDB code http://www.rcsb.org/pdb/search/structidSearch.do?structureId=3KG1, 10% identity with OAC) , Mycobacterium tuberculosis Rv0793 (PDB code http://www.rcsb.org/pdb/search/structidSearch.do?structureId=1Y0H, 7% identity with OAC) , MLMI (PDB code http://www.rcsb.org/pdb/search/structidSearch.do?structureId=3HDS, 13% identity with OAC) , ActVA‐Orf6 (PDB code http://www.rcsb.org/pdb/search/structidSearch.do?structureId=1N5S, 15% identity with OAC) , Pseudomonas aeruginosa PA3566 (PDB code http://www.rcsb.org/pdb/search/structidSearch.do?structureId=1X7V, 7% identity with OAC) and TcmI (PDB code http://www.rcsb.org/pdb/search/structidSearch.do?structureId=1TUW, 17% identity with OAC) , respectively (Figs and ).…”

Section: Resultsmentioning

confidence: 99%

Structural basis for olivetolic acid formation by a polyketide cyclase from Cannabis sativa

et al. 2016

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In polyketide biosynthesis, ring formation is one of the key diversification steps. Olivetolic acid cyclase (OAC) from Cannabis sativa, involved in cannabinoid biosynthesis, is the only known plant polyketide cyclase. In addition, it is the only functionally characterized plant a+b barrel (DABB) protein that catalyzes the C2-C7 aldol cyclization of the linear pentyl tetra-b-ketide CoA as the substrate, to generate olivetolic acid (OA). Herein, we solved the OAC apo and OAC-OA complex binary crystal structures at 1.32 and 1.70A resolutions, respectively. The crystal structures revealed that the enzyme indeed belongs to the DABB superfamily, as previously proposed, and possesses a unique active-site cavity containing the pentyl-binding hydrophobic pocket and the polyketide binding site, which have never been observed among the functionally and structurally characterized bacterial polyketide cyclases. Furthermore, site-directed mutagenesis studies indicated that Tyr72 and His78 function as acid/base catalysts at the catalytic center. Structural and/or functional studies of OAC suggested that the enzyme lacks thioesterase and aromatase activities. These observations demonstrated that OAC employs unique catalytic machinery utilizing acid/base catalytic chemistry for the formation of the precursor of OA. The structural and functional insights obtained in this work thus provide the foundation for analyses of the plant polyketide cyclases that will be discovered in the future.

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“…They all act independently of co-factors, prosthetic group and metals. The substrates vary greatly in size, which is reflected in the size of the active site cavities of the enzymes (43). The fact that substrates such as heme groups (porphyrin structure), anthracycline (four aromatic rings), and actinorhodin (three aromatic rings) can be accommodated by this type of protein fold makes it likely that AurZ can bind YWA1 in a similar fashion.…”

Section: Discussionmentioning

confidence: 99%

Two Novel Classes of Enzymes Are Required for the Biosynthesis of Aurofusarin in Fusarium graminearum

Frandsen

Schütt

Lund

et al. 2011

Journal of Biological Chemistry

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Previous studies have reported the functional characterization of 9 out of 11 genes found in the gene cluster responsible for biosynthesis of the polyketide pigment aurofusarin in Fusarium graminearum. Here we reanalyze the function of a putative aurofusarin pump (AurT) and the two remaining orphan genes, aurZ and aurS. Targeted gene replacement of aurZ resulted in the discovery that the compound YWA1, rather than nor-rubrofusarin, is the primary product of F. graminearum polyketide synthase 12 (FgPKS12). AurZ is the first representative of a novel class of dehydratases that act on hydroxylated ␥-pyrones. Replacement of the aurS gene resulted in accumulation of rubrofusarin, an intermediate that also accumulates when the GIP1, aurF, or aurO genes in the aurofusarin cluster are deleted. Based on the shared phenotype and predicted subcellular localization, we propose that AurS is a member of an extracellular enzyme complex (GIP1-AurF-AurO-AurS) responsible for converting rubrofusarin into aurofusarin. This implies that rubrofusarin, rather than aurofusarin, is pumped across the plasma membrane. Replacement of the putative aurofusarin pump aurT increased the rubrofusarin-to-aurofusarin ratio, supporting that rubrofusarin is normally pumped across the plasma membrane. These results provide functional information on two novel classes of proteins and their contribution to polyketide pigment biosynthesis.The plant pathogenic fungus Fusarium graminearum (teleomorph Gibberella zeae) is capable of producing a plethora of secondary metabolites, many of which belong to the polyketide class of compounds, such as the mycotoxins zearalenone, fusarin C, and aurofusarin (1). The F. graminearum genome encodes 15 polyketide synthases (PKS) 2 (2, 3), of which FgPKS12 is the progenitor of nor-rubrofusarin/rubrofusarin/aurofusarin (4, 5), FgPKS4 and FgPKS13 are the progenitors of zearalenone (1, 6), FgPKS10 is the progenitor of fusarin C (7, 8), and FgPKS3 is the progenitor of an uncharacterized black/purple perithecial pigment (8). PKS genes are typically found in gene clusters that comprise genes encoding transcription factor, transporters, and tailoring enzymes required for biosynthesis of the final product (1, 4 -6). In addition to genes encoding well characterized classes of enzymes, the clusters typically also include orphan genes that encode proteins annotated as "hypothetical" or "conserved hypothetical proteins," signifying that no similarity to any previously characterized protein has been found. In the case of F. graminearum, more than half of all gene models found in the Fusarium graminearum Genome Database from the Munich Information Center for Protein Sequences (MIPS) fall into either of these two categories (9).Functional characterization of hypothetical proteins is challenging, but those belonging to secondary metabolite gene clusters provide a unique opportunity because their location suggests a function in the respective biosynthetic pathways. The PKS12 gene cluster in F. graminearum, which is responsible for biosyn...

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The crystal structure of Rv0793, a hypothetical monooxygenase from M.␣tuberculosis

Cited by 15 publications

References 34 publications

Structural and functional characterization of Rv0792c from Mycobacterium tuberculosis: identifying small molecule inhibitors against GntR protein

Structural and functional characterization of Rv0792c from Mycobacterium tuberculosis: identifying small molecule inhibitors against GntR protein

Structural basis for olivetolic acid formation by a polyketide cyclase from Cannabis sativa

Two Novel Classes of Enzymes Are Required for the Biosynthesis of Aurofusarin in Fusarium graminearum

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