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Sample records for a32 modular polyketide

  1. Iterative Polyketide Biosynthesis by Modular Polyketide Synthases in Bacteria

    PubMed Central

    Chen, Haotong; Du, Liangcheng

    2015-01-01

    Modular polyketide synthases (type I PKSs) in bacteria are responsible for synthesizing a significant percentage of bioactive natural products. This group of synthases has a characteristic modular organization, and each module within a PKS carries out one cycle of polyketide chain elongation; thus each module is “non-iterative” in function. It was possible to predict the basic structure of a polyketide product from the module organization of the PKSs, since there generally existed a co-linearity between the number of modules and the number of chain elongations. However, more and more bacterial modular PKSs fail to conform to the “canonical rules”, and a particularly noteworthy group of non-canonical PKSs is the bacterial iterative type I PKSs. This review covers recent examples of iteratively-used modular PKSs in bacteria. These non-canonical PKSs give rise to a large array of natural products with impressive structural diversity. The molecular mechanism behind the iterations is often unclear, presenting a new challenge to the rational engineering of these PKSs with the goal of generating new natural products. Structural elucidation of these synthase complexes and better understanding of potential PKS-PKS interactions as well as PKS-substrate recognition may provide new prospects and inspirations for the discovery and engineering of new bioactive polyketides. PMID:26549236

  2. Structure of a modular polyketide synthase

    PubMed Central

    Dutta, Somnath; Whicher, Jonathan R.; Hansen, Douglas A.; Hale, Wendi A.; Chemler, Joseph A.; Congdon, Grady R.; Narayan, Alison R.; Håkansson, Kristina; Sherman, David H.; Smith, Janet L.

    2014-01-01

    Polyketide natural products constitute a broad class of compounds with diverse structural features and biological activities. Their biosynthetic machinery, represented by type I polyketide synthases, has an architecture in which successive modules catalyze two-carbon linear extensions and keto group processing reactions on intermediates covalently tethered to carrier domains. We employed electron cryo-microscopy to visualize a full-length module and determine sub-nanometer resolution 3D reconstructions that revealed an unexpectedly different architecture compared to the homologous dimeric mammalian fatty acid synthase. A single reaction chamber provides access to all catalytic sites for the intra-module carrier domain. In contrast, the carrier from the preceding module uses a separate entrance outside the reaction chamber to deliver the upstream polyketide intermediate for subsequent extension and modification. This study reveals for the first time the structural basis for both intra-module and inter-module substrate transfer in polyketide synthases, and establishes a new model for molecular dissection of these multifunctional enzyme systems. PMID:24965652

  3. Evolution of Efficient Modular Polyketide Synthases by Homologous Recombination

    PubMed Central

    Chemler, Joseph A.; Tripathi, Ashootosh; Hansen, Douglas A.; O'Neil-Johnson, Mark; Williams, Russell B.; Starks, Courtney; Park, Sung Ryeol; Sherman, David H.

    2015-01-01

    The structural scaffolds of many complex natural products are produced by multifunctional type I polyketide synthase (PKS) enzymes that operate as bio-synthetic assembly lines. The modular nature of these megaenzymes presents an opportunity to construct custom biocatalysts built in a lego-like fashion by inserting, deleting, or exchanging native or foreign domains to produce targeted variants of natural polyketides. However, previously engineered PKS enzymes are often impaired resulting in limited production compared to native systems. Here, we show a versatile method for generating and identifying functional chimeric PKS enzymes for synthesizing custom macrolactones and macrolides. PKS genes from the pikromycin and erythromycin pathways were hybridized in Saccharomyces cerevisiae to generate hybrid libraries. We used a 96-well plate format for plasmid purification, transformations, sequencing, protein expression, in vitro reactions and analysis of metabolite formation. Active chimeric enzymes were identified with new functionality. Streptomyces venezuelae strains that expressed these PKS chimeras were capable of producing engineered macrolactones. Furthermore, a macrolactone generated from selected PKS chimeras was fully functionalized into a novel macrolide analogue. This method permits the engineering of PKS pathways as modular building blocks for the production of new antibiotic-like molecules. PMID:26230368

  4. MOLECULAR DYNAMICS STUDIES OF MODULAR POLYKETIDE SYNTHASE KETOREDUCTASE STEREOSPECIFICITY

    PubMed Central

    Mugnai, Mauro L.; Shi, Yue; Keatinge-Clay, Adrian T.; Elber, Ron

    2015-01-01

    Ketoreductases (KRs) from modular polyketide synthases (PKSs) can perform stereospecific catalysis, selecting a polyketide with a D-α or an L-α-methyl substituent for NADPH-mediated reduction. In this report Molecular Dynamics (MD) simulations were performed to investigate the interactions that control stereospecificity. We studied the A1-type KR from the second module of the amphotericin PKS (A1), which is known to be stereospecific for a D-α-methyl-substituted diketide substrate (dkD). MD simulations of two ternary complexes comprised of the enzyme, NADPH, and either the correct substrate, dkD, or its enantiomer (dkL) were performed. The coordinates for the A1/NADPH binary complex were obtained from a crystal structure [Zheng, J. T. et al. (2010) Structure 18, 913–922], and substrates were modeled in the binding pocket in conformations appropriate for reduction. Simulations were intended to reproduce the initial weak binding of the polyketide substrate to the enzyme. Long (tens of nanoseconds) MD simulations show that the correct substrate is retained in a conformation closer to the reactive configuration. Many short (up to a nanosecond) MD runs starting from the initial structures display evidence that Q364, three residues N-terminal to the catalytic tyrosine, forms a hydrogen bond to the incorrect dkL substrate to yield an unreactive conformation that is more favorable than the reactive configuration. This interaction is not as strong for dkD, as the D-α-methyl substituent is positioned between the glutamine and the reactive site. This result correlates with experimental findings [Zheng, J. T. et al. (2010) Structure 18, 913–922] in which a Q364H mutant was observed to lose stereospecificity. PMID:25835227

  5. A close look at a ketosynthase from a trans-acyltransferase modular polyketide synthase

    PubMed Central

    Gay, Darren C.; Gay, Glen; Axelrod, Abram J.; Jenner, Matthew; Kohlhaas, Christoph; Kampa, Annette; Oldham, Neil J.; Piel, Jörn; Keatinge-Clay, Adrian T.

    2014-01-01

    SUMMARY The recently discovered trans-acyltransferase modular polyketide synthases catalyze the biosynthesis of a wide range of bioactive natural products in bacteria. Here we report the structure of the second ketosynthase from the bacillaene trans-acyltransferase polyketide synthase. This 1.95 Å-resolution structure provides the highest resolution view available of a modular polyketide synthase ketosynthase and reveals a flanking subdomain that is homologous to an ordered linker in cis-acyltransferase modular polyketide synthases. The structure of the cysteine-to-serine mutant of the ketosynthase acylated by its natural substrate provides high-resolution details of how a native polyketide intermediate is bound and helps explain the basis of ketosynthase substrate specificity. The substrate range of the ketosynthase was further investigated by mass spectrometry. PMID:24508341

  6. Engineering modular polyketide synthases for production of biofuels and industrial chemicals.

    PubMed

    Cai, Wenlong; Zhang, Wenjun

    2017-09-22

    Polyketide synthases (PKSs) are one of the most profound biosynthetic factories for producing polyketides with diverse structures and biological activities. These enzymes have been historically studied and engineered to make un-natural polyketides for drug discovery, and have also recently been explored for synthesizing biofuels and industrial chemicals due to their versatility and customizability. Here, we review recent advances in the mechanistic understanding and engineering of modular PKSs for producing polyketide-derived chemicals, and provide perspectives on this relatively new application of PKSs. Copyright © 2017 Elsevier Ltd. All rights reserved.

  7. Heterologous production of polyketides by modular type I polyketide synthases in Escherichia coli

    SciTech Connect

    Yuzawa, S; Kim, W; Katz, L; Keasling, JD

    2012-10-01

    Heterologous production of polyketide compounds, an important class of natural products with complex chemical structures, was first demonstrated with Streptomyces parvulus in 1984. Although Streptomyces strains are good first options for heterologous polyketide biosynthesis, their slow growth kinetics prompt other hosts to also be considered. Escherichia coli provides key elements of an ideal host in terms of the growth rate, culture conditions, and available recombinant DNA tools. Here we review the current status and potential for metabolic engineering of polyketides in E. coli.

  8. Alteration of Polyketide Stereochemistry from anti to syn by a Ketoreductase Domain Exchange in a Type I Modular Polyketide Synthase Subunit.

    PubMed

    Eng, Clara H; Yuzawa, Satoshi; Wang, George; Baidoo, Edward E K; Katz, Leonard; Keasling, Jay D

    2016-03-29

    Polyketide natural products have broad applications in medicine. Exploiting the modular nature of polyketide synthases to alter stereospecificity is an attractive strategy for obtaining natural product analogues with altered pharmaceutical properties. We demonstrate that by retaining a dimerization element present in LipPks1+TE, we are able to use a ketoreductase domain exchange to alter α-methyl group stereochemistry with unprecedented retention of activity and simultaneously achieve a novel alteration of polyketide product stereochemistry from anti to syn. The substrate promiscuity of LipPks1+TE further provided a unique opportunity to investigate the substrate dependence of ketoreductase activity in a polyketide synthase module context.

  9. Structural and functional analysis of the loading acyltransferase from avermectin modular polyketide synthase.

    PubMed

    Wang, Fen; Wang, Yanjie; Ji, Junjie; Zhou, Zhan; Yu, Jingkai; Zhu, Hua; Su, Zhiguo; Zhang, Lixin; Zheng, Jianting

    2015-04-17

    The loading acyltransferase (AT) domains of modular polyketide synthases (PKSs) control the choice of starter units incorporated into polyketides and are therefore attractive targets for the engineering of modular PKSs. Here, we report the structural and biochemical characterizations of the loading AT from avermectin modular PKS, which accepts more than 40 carboxylic acids as alternative starter units for the biosynthesis of a series of congeners. This first structural analysis of loading ATs from modular PKSs revealed the molecular basis for the relaxed substrate specificity. Residues important for substrate binding and discrimination were predicted by modeling a substrate into the active site. A mutant with altered specificity toward a panel of synthetic substrate mimics was generated by site-directed mutagenesis of the active site residues. The hydrolysis of the N-acetylcysteamine thioesters of racemic 2-methylbutyric acid confirmed the stereospecificity of the avermectin loading AT for an S configuration at the C-2 position of the substrate. Together, these results set the stage for region-specific modification of polyketides through active site engineering of loading AT domains of modular PKSs.

  10. Alteration of Polyketide Stereochemistry from anti to syn by a Ketoreductase Domain Exchange in a Type I Modular Polyketide Synthase Subunit

    SciTech Connect

    Eng, Clara H.; Yuzawa, Satoshi; Wang, George; Baidoo, Edward E. K.; Katz, Leonard; Keasling, Jay D.

    2016-03-29

    Polyketide natural products have broad applications in medicine. Exploiting the modular nature of polyketide synthases to alter stereospecificity is an attractive strategy for obtaining natural product analogues with altered pharmaceutical properties. The purpose of research is to demonstrate that by retaining a dimerization element present in LipPks1+TE, we are able to use a ketoreductase domain exchange to alter α-methyl group stereochemistry with unprecedented retention of activity and simultaneously achieve a novel alteration of polyketide product stereochemistry from anti to syn. The substrate promiscuity of LipPks1+TE further provided a unique opportunity to investigate the substrate dependence of ketoreductase activity in a polyketide synthase module context.

  11. Substrate structure-activity relationships guide rational engineering of modular polyketide synthase ketoreductases.

    PubMed

    Bailey, Constance B; Pasman, Marjolein E; Keatinge-Clay, Adrian T

    2016-01-14

    Modular polyketide synthase ketoreductases can set two chiral centers through a single reduction. To probe the basis of stereocontrol, a structure-activity relationship study was performed with three α-methyl, β-ketothioester substrates and four ketoreductases. Since interactions with the β-ketoacyl moiety were found to be most critical, residues implicated in contacting this moiety were mutated. Two mutations were sufficient to completely reverse the stereoselectivity of the model ketoreductase EryKR1, converting it from an enzyme that generates (2S,3R)-products into one that yields (2S,3S)-products.

  12. Biochemical and Structural Basis for Controlling Chemical Modularity in Fungal Polyketide Biosynthesis

    SciTech Connect

    Winter, Jaclyn M.; Cascio, Duilio; Dietrich, David; Sato, Michio; Watanabe, Kenji; Sawaya, Michael R.; Vederas, John C.; Tang, Yi

    2015-07-14

    Modular collaboration between iterative fungal polyketide synthases (IPKSs) is an important mechanism for generating structural diversity of polyketide natural products. Inter-PKS communication and substrate channeling are controlled in large by the starter unit acyl carrier protein transacylase (SAT) domain found in the accepting IPKS module. Here in this study, we reconstituted the modular biosynthesis of the benzaldehyde core of the chaetoviridin and chaetomugilin azaphilone natural products using the IPKSs CazF and CazM. Our studies revealed a critical role of CazM’s SAT domain in selectively transferring a highly reduced triketide product from CazF. In contrast, a more oxidized triketide that is also produced by CazF and required in later stages of biosynthesis of the final product is not recognized by the SAT domain. The structural basis for the acyl unit selectivity was uncovered by the first X-ray structure of a fungal SAT domain, highlighted by a covalent hexanoyl thioester intermediate in the SAT active site. Finally, the crystal structure of SAT domain will enable protein engineering efforts aimed at mixing and matching different IPKS modules for the biosynthesis of new compounds.

  13. Biochemical and Structural Basis for Controlling Chemical Modularity in Fungal Polyketide Biosynthesis

    DOE PAGES

    Winter, Jaclyn M.; Cascio, Duilio; Dietrich, David; ...

    2015-07-14

    Modular collaboration between iterative fungal polyketide synthases (IPKSs) is an important mechanism for generating structural diversity of polyketide natural products. Inter-PKS communication and substrate channeling are controlled in large by the starter unit acyl carrier protein transacylase (SAT) domain found in the accepting IPKS module. Here in this study, we reconstituted the modular biosynthesis of the benzaldehyde core of the chaetoviridin and chaetomugilin azaphilone natural products using the IPKSs CazF and CazM. Our studies revealed a critical role of CazM’s SAT domain in selectively transferring a highly reduced triketide product from CazF. In contrast, a more oxidized triketide that ismore » also produced by CazF and required in later stages of biosynthesis of the final product is not recognized by the SAT domain. The structural basis for the acyl unit selectivity was uncovered by the first X-ray structure of a fungal SAT domain, highlighted by a covalent hexanoyl thioester intermediate in the SAT active site. Finally, the crystal structure of SAT domain will enable protein engineering efforts aimed at mixing and matching different IPKS modules for the biosynthesis of new compounds.« less

  14. Loss of co-linearity by modular polyketide synthases: a mechanism for the evolution of chemical diversity.

    PubMed

    Moss, Steven J; Martin, Christine J; Wilkinson, Barrie

    2004-10-01

    Modular polyketide synthases biosynthesise natural products through successive Claisen-type condensations, where one module is responsible for one round of chain extension. This review describes recent findings where this rule of co-linearity is broken, either by one module being bypassed (skipping) or through one module being used for multiple chain extension events (stuttering).

  15. Targeting modular polyketide synthases with iteratively acting acyltransferases from metagenomes of uncultured bacterial consortia.

    PubMed

    Piel, Jörn; Hui, Dequan; Fusetani, Nobuhiro; Matsunaga, Shigeki

    2004-09-01

    Bacterial type I polyketide synthases (PKSs) produce a wide range of biomedically important secondary metabolites. These enzymes possess a modular structure that can be genetically re-engineered to yield novel drug candidates not found in nature. Recently, we have reported the putative pederin PKS from an uncultured bacterial symbiont of Paederus fuscipes beetles. It belongs to an architecturally unusual PKS group, the members of which contain iteratively acting acyltransferases that are not integrated into the PKS modules but are encoded by isolated genes. As these systems are rare, often contain additional unusual features and are of smaller size than regular PKSs, the development of a method for the targeted isolation of new group members would be of great interest. Here, we present a phylogenetic approach to identify these systems rapidly in highly complex metagenomic DNA samples. To demonstrate its practical value, we located two pederin-type PKS systems putatively involved in the biosynthesis of antitumour polyketides in the metagenomic DNA of beetles, sponges and their uncultivated bacterial symbionts.

  16. Bio-based production of fuels and industrial chemicals by repurposing antibiotic-producing type I modular polyketide synthases: opportunities and challenges.

    PubMed

    Yuzawa, Satoshi; Keasling, Jay D; Katz, Leonard

    2017-04-01

    Complex polyketides comprise a large number of natural products that have broad application in medicine and agriculture. They are produced in bacteria and fungi from large enzyme complexes named type I modular polyketide synthases (PKSs) that are composed of multifunctional polypeptides containing discrete enzymatic domains organized into modules. The modular nature of PKSs has enabled a multitude of efforts to engineer the PKS genes to produce novel polyketides of predicted structure. We have repurposed PKSs to produce a number of short-chain mono- and di-carboxylic acids and ketones that could have applications as fuels or industrial chemicals.

  17. Bio-based production of fuels and industrial chemicals by repurposing antibiotic-producing type I modular polyketide synthases: opportunities and challenges

    SciTech Connect

    Yuzawa, Satoshi; Keasling, Jay D.; Katz, Leonard

    2016-11-16

    Complex polyketides comprise a large number of natural products that have broad application in medicine and agriculture. They are produced in bacteria and fungi from large enzyme complexes named type I modular polyketide synthases (PKSs) that are composed of multifunctional polypeptides containing discrete enzymatic domains organized into modules. The modular nature of PKSs has enabled a multitude of efforts to engineer the PKS genes to produce novel polyketides of predicted structure. Finally, we have repurposed PKSs to produce a number of short-chain mono- and di-carboxylic acids and ketones that could have applications as fuels or industrial chemicals.

  18. Polyketide genes in the marine sponge Plakortis simplex: a new group of mono-modular type I polyketide synthases from sponge symbionts

    PubMed Central

    Della Sala, Gerardo; Hochmuth, Thomas; Costantino, Valeria; Teta, Roberta; Gerwick, William; Gerwick, Lena; Piel, Jörn; Mangoni, Alfonso

    2013-01-01

    Summary Sponge symbionts are a largely unexplored source of new and unusual metabolic pathways. Insights into the distribution and function of metabolic genes of sponge symbionts are crucial to dissect and exploit their biotechnological potential. Screening of the metagenome of the marine sponge Plakortis simplex led to the discovery of the swf family, a new group of mono-modular type I polyketide synthase/fatty acid synthase (PKS/FAS) specifically associated with sponge symbionts. Two different examples of the swf cluster were present in the metagenome of P. simplex. A third example of the cluster is present in the previously sequenced genome of a poribacterium from the sponge Aplysina aerophoba but was formerly considered orthologous to the wcb/rkp cluster. The swf cluster was also found in six additional species of sponges. Therefore, the swf cluster represents the second group of mono-modular PKS, after the supA family, to be widespread in marine sponges. The putative swf operon consists of swfA (type I PKS/FAS), swfB (reductase and sulphotransferase domains) and swfC (radical S-adenosylmethionine, or radical SAM). Activation of the acyl carrier protein (ACP) domain of the SwfA protein to its holo-form by co-expression with Svp is the first functional proof of swf type genes in marine sponges. However, the precise biosynthetic role of the swf clusters remains unknown. PMID:24249289

  19. Generation of multiple bioactive macrolides by hybrid modular polyketide synthases in Streptomyces venezuelae.

    PubMed

    Yoon, Yeo Joon; Beck, Brian J; Kim, Beom Seok; Kang, Han Young; Reynolds, Kevin A; Sherman, David H

    2002-02-01

    The plasmid-based replacement of the multifunctional protein subunits of the pikromycin PKS in S. venezuelae by the corresponding subunits from heterologous modular PKSs resulted in recombinant strains that produce both 12- and 14-membered ring macrolactones with predicted structural alterations. In all cases, novel macrolactones were produced and further modified by the DesVII glycosyltransferase and PikC hydroxylase, leading to biologically active macrolide structures. These results demonstrate that hybrid PKSs in S. venezuelae can produce a multiplicity of new macrolactones that are modified further by the highly flexible DesVII glycosyltransferase and PikC hydroxylase tailoring enzymes. This work demonstrates the unique capacity of the S. venezuelae pikromycin pathway to expand the toolbox of combinatorial biosynthesis and to accelerate the creation of novel biologically active natural products.

  20. Creating polyketide diversity through genetic engineering.

    PubMed

    Kealey, James T

    2003-01-01

    Modular polyketide synthases (PKS) are large multifunctional enzymes that synthesize complex polyketides, a therapeutically important class of natural products. The linear order and composition of catalytic sites that comprise the PKS represent a "code" that determines the identity of the polyketide product. By re-programming the PKS through genetic engineering, it is possible to alter the code in a predictable manner to create specific structural modifications of polyketides and to produce new libraries of these natural products.

  1. Comprehensive in Vitro Analysis of Acyltransferase Domain Exchanges in Modular Polyketide Synthases and Its Application for Short-Chain Ketone Production.

    PubMed

    Yuzawa, Satoshi; Deng, Kai; Wang, George; Baidoo, Edward E K; Northen, Trent R; Adams, Paul D; Katz, Leonard; Keasling, Jay D

    2017-01-20

    Type I modular polyketide synthases (PKSs) are polymerases that utilize acyl-CoAs as substrates. Each polyketide elongation reaction is catalyzed by a set of protein domains called a module. Each module usually contains an acyltransferase (AT) domain, which determines the specific acyl-CoA incorporated into each condensation reaction. Although a successful exchange of individual AT domains can lead to the biosynthesis of a large variety of novel compounds, hybrid PKS modules often show significantly decreased activities. Using monomodular PKSs as models, we have systematically analyzed the segments of AT domains and associated linkers in AT exchanges in vitro and have identified the boundaries within a module that can be used to exchange AT domains while maintaining protein stability and enzyme activity. Importantly, the optimized domain boundary is highly conserved, which facilitates AT domain replacements in most type I PKS modules. To further demonstrate the utility of the optimized AT domain boundary, we have constructed hybrid PKSs to produce industrially important short-chain ketones. Our in vitro and in vivo analysis demonstrated production of predicted ketones without significant loss of activities of the hybrid enzymes. These results greatly enhance the mechanistic understanding of PKS modules and prove the benefit of using engineered PKSs as a synthetic biology tool for chemical production.

  2. Comprehensive in Vitro Analysis of Acyltransferase Domain Exchanges in Modular Polyketide Synthases and Its Application for Short-Chain Ketone Production

    SciTech Connect

    Yuzawa, Satoshi; Deng, Kai; Wang, George; Baidoo, Edward E. K.; Northen, Trent R.; Adams, Paul D.; Katz, Leonard; Keasling, Jay D.

    2016-08-22

    Type I modular polyketide synthases (PKSs) are polymerases that utilize acyl-CoAs as substrates. Each polyketide elongation reaction is catalyzed by a set of protein domains called a module. Each module usually contains an acyltransferase (AT) domain, which determines the specific acyl-CoA incorporated into each condensation reaction. Although a successful exchange of individual AT domains can lead to the biosynthesis of a large variety of novel compounds, hybrid PKS modules often show significantly decreased activities. Using monomodular PKSs as models, we have systematically analyzed in this paper the segments of AT domains and associated linkers in AT exchanges in vitro and have identified the boundaries within a module that can be used to exchange AT domains while maintaining protein stability and enzyme activity. Importantly, the optimized domain boundary is highly conserved, which facilitates AT domain replacements in most type I PKS modules. To further demonstrate the utility of the optimized AT domain boundary, we have constructed hybrid PKSs to produce industrially important short-chain ketones. Our in vitro and in vivo analysis demonstrated production of predicted ketones without significant loss of activities of the hybrid enzymes. Finally, these results greatly enhance the mechanistic understanding of PKS modules and prove the benefit of using engineered PKSs as a synthetic biology tool for chemical production.

  3. Dissecting complex polyketide biosynthesis

    PubMed Central

    Caffrey, Patrick

    2012-01-01

    Numerous bioactive natural products are synthesised by modular polyketide synthases. These compounds can be made in high yield by native multienzyme assembly lines. However, formation of analogues by genetically engineered systems is often considerably less efficient. Biochemical studies on intact polyketide synthase proteins have amassed a body of knowledge that is substantial but still incomplete. Recently, the constituent enzymes have been structurally characterised as discrete domains or didomains. These recombinant proteins have been used to reconstitute single extension cycles in vitro. This has given further insights into how the final stereochemistry of chiral centres in polyketides is determined. In addition, this approach has revealed how domains co-operate to ensure efficient transfer of growing intermediates along the assembly line. This work is leading towards more effective re-programming of these enzymes for use in synthesis of new medicinal compounds. PMID:24688670

  4. Modeling holo-ACP:DH and holo-ACP:KR complexes of modular polyketide synthases: a docking and molecular dynamics study.

    PubMed

    Anand, Swadha; Mohanty, Debasisa

    2012-05-28

    Modular polyketide synthases are multifunctional megasynthases which biosynthesize a variety of secondary metabolites using various combinations of dehydratase (DH), ketoreductase (KR) and enoyl-reductase (ER) domains. During the catalysis of various reductive steps these domains act on a substrate moiety which is covalently attached to the phosphopantetheine (P-pant) group of the holo-Acyl Carrier Protein (holo-ACP) domain, thus necessitating the formation of holo-ACP:DH and holo-ACP:KR complexes. Even though three dimensional structures are available for DH, KR and ACP domains, no structures are available for DH or KR domains in complex with ACP or substrate moieties. Since Ser of holo-ACP is covalently attached to a large phosphopantetheine group, obtaining complexes involving holo-ACP by standard protein-protein docking has been a difficult task. We have modeled the holo-ACP:DH and holo-ACP:KR complexes for identifying specific residues on DH and KR domains which are involved in interaction with ACP, phosphopantetheine and substrate moiety. A novel combination of protein-protein and protein-ligand docking has been used to first model complexes involving apo-ACP and then dock the phosphopantetheine and substrate moieties using covalent connectivity between ACP, phosphopantetheine and substrate moiety as constraints. The holo-ACP:DH and holo-ACP:KR complexes obtained from docking have been further refined by restraint free explicit solvent MD simulations to incorporate effects of ligand and receptor flexibilities. The results from 50 ns MD simulations reveal that substrate enters into a deep tunnel in DH domain while in case of KR domain the substrate binds a shallow surface exposed cavity. Interestingly, in case of DH domain the predicted binding site overlapped with the binding site in the inhibitor bound crystal structure of FabZ, the DH domain from E.Coli FAS. In case of KR domain, the substrate binding site identified by our simulations was in proximity of

  5. Modeling holo-ACP:DH and holo-ACP:KR complexes of modular polyketide synthases: a docking and molecular dynamics study

    PubMed Central

    2012-01-01

    Background Modular polyketide synthases are multifunctional megasynthases which biosynthesize a variety of secondary metabolites using various combinations of dehydratase (DH), ketoreductase (KR) and enoyl-reductase (ER) domains. During the catalysis of various reductive steps these domains act on a substrate moiety which is covalently attached to the phosphopantetheine (P-pant) group of the holo-Acyl Carrier Protein (holo-ACP) domain, thus necessitating the formation of holo-ACP:DH and holo-ACP:KR complexes. Even though three dimensional structures are available for DH, KR and ACP domains, no structures are available for DH or KR domains in complex with ACP or substrate moieties. Since Ser of holo-ACP is covalently attached to a large phosphopantetheine group, obtaining complexes involving holo-ACP by standard protein-protein docking has been a difficult task. Results We have modeled the holo-ACP:DH and holo-ACP:KR complexes for identifying specific residues on DH and KR domains which are involved in interaction with ACP, phosphopantetheine and substrate moiety. A novel combination of protein-protein and protein-ligand docking has been used to first model complexes involving apo-ACP and then dock the phosphopantetheine and substrate moieties using covalent connectivity between ACP, phosphopantetheine and substrate moiety as constraints. The holo-ACP:DH and holo-ACP:KR complexes obtained from docking have been further refined by restraint free explicit solvent MD simulations to incorporate effects of ligand and receptor flexibilities. The results from 50 ns MD simulations reveal that substrate enters into a deep tunnel in DH domain while in case of KR domain the substrate binds a shallow surface exposed cavity. Interestingly, in case of DH domain the predicted binding site overlapped with the binding site in the inhibitor bound crystal structure of FabZ, the DH domain from E.Coli FAS. In case of KR domain, the substrate binding site identified by our simulations

  6. Genetic engineering to produce polyketide analogues.

    PubMed

    Reeves, Christopher D; Rodriguez, Eduardo

    2009-01-01

    Polyketides are pharmaceutically important and structurally diverse natural products. Creating analogues for drug development can be done with chemistry, but this is generally restricted to a few accessible functional groups. Analogues can also be made by genetic engineering, which is particularly effective for polyketides synthesized by a modular polyketide synthase (PKS). Such a PKS displays colinearity, which means that the structural features along the polyketide chain are determined by the catalytic specificities in corresponding modules along a molecular assembly line. The assembly line can be genetically engineered through addition, deletion, or mutation of catalytic domains or the reorganization of whole modules. Chemically synthesized precursors also can be fed to engineered assembly lines to further expand the repertoire of analogues. These various methods are discussed with an aim of providing a guide to the strategies most likely to succeed in a given circumstance. Recent information that could be relevant to future polyketide engineering projects is also discussed.

  7. The Assembly Line Enzymology of Polyketide Biosynthesis.

    PubMed

    Till, Marisa; Race, Paul R

    2016-01-01

    Polyketides are a structurally and functionally diverse family of bioactive natural products that have found widespread application as pharmaceuticals, agrochemicals, and veterinary medicines. In bacteria complex polyketides are biosynthesized by giant multifunctional megaenzymes, termed modular polyketide synthases (PKSs), which construct their products in a highly coordinated assembly line-like fashion from a pool of simple precursor substrates. Not only is the multifaceted enzymology of PKSs a fascinating target for study, but it also presents considerable opportunities for the reengineering of these systems affording access to functionally optimized unnatural natural products. Here we provide an introductory primer to modular polyketide synthase structure and function, and highlight recent advances in the characterization and exploitation of these systems.

  8. Bioinspired iterative synthesis of polyketides

    PubMed Central

    Zheng, Kuan; Xie, Changmin; Hong, Ran

    2015-01-01

    Diverse array of biopolymers and second metabolites (particularly polyketide natural products) has been manufactured in nature through an enzymatic iterative assembly of simple building blocks. Inspired by this strategy, molecules with inherent modularity can be efficiently synthesized by repeated succession of similar reaction sequences. This privileged strategy has been widely adopted in synthetic supramolecular chemistry. Its value also has been reorganized in natural product synthesis. A brief overview of this approach is given with a particular emphasis on the total synthesis of polyol-embedded polyketides, a class of vastly diverse structures and biologically significant natural products. This viewpoint also illustrates the limits of known individual modules in terms of diastereoselectivity and enantioselectivity. More efficient and practical iterative strategies are anticipated to emerge in the future development. PMID:26052510

  9. Role of Modular Polyketide Synthases in the Production of Polyether Ladder Compounds in Ciguatoxin-Producing Gambierdiscus polynesiensis and G. excentricus (Dinophyceae).

    PubMed

    Kohli, Gurjeet S; Campbell, Katrina; John, Uwe; Smith, Kirsty F; Fraga, Santiago; Rhodes, Lesley L; Murray, Shauna A

    2017-02-17

    Gambierdiscus, a benthic dinoflagellate, produces ciguatoxins that cause the human illness Ciguatera. Ciguatoxins are polyether ladder compounds that have a polyketide origin, indicating that polyketide synthases (PKS) are involved in their production. We sequenced transcriptomes of Gambierdiscus excentricus and Gambierdiscus polynesiensis and found 264 contigs encoding single domain ketoacyl synthases (KS; G. excentricus: 106, G. polynesiensis: 143) and ketoreductases (KR; G. excentricus: 7, G. polynesiensis: 8) with sequence similarity to type I PKSs, as reported in other dinoflagellates. In addition, 24 contigs (G. excentricus: 3, G. polynesiensis: 21) encoding multiple PKS domains (forming typical type I PKSs modules) were found. The proposed structure produced by one of these megasynthases resembles a partial carbon backbone of a polyether ladder compound. Seventeen contigs encoding single domain KS, KR, s-malonyltransacylase, dehydratase and enoyl reductase with sequence similarity to type II fatty acid synthases (FAS) in plants were found. Type I PKS and type II FAS genes were distinguished based on the arrangement of domains on the contigs and their sequence similarity and phylogenetic clustering with known PKS/FAS genes in other organisms. This differentiation of PKS and FAS pathways in Gambierdiscus is important, as it will facilitate approaches to investigating toxin biosynthesis pathways in dinoflagellates.

  10. Polyketide stereocontrol: a study in chemical biology

    PubMed Central

    2017-01-01

    The biosynthesis of reduced polyketides in bacteria by modular polyketide synthases (PKSs) proceeds with exquisite stereocontrol. As the stereochemistry is intimately linked to the strong bioactivity of these molecules, the origins of stereochemical control are of significant interest in attempts to create derivatives of these compounds by genetic engineering. In this review, we discuss the current state of knowledge regarding this key aspect of the biosynthetic pathways. Given that much of this information has been obtained using chemical biology tools, work in this area serves as a showcase for the power of this approach to provide answers to fundamental biological questions. PMID:28326145

  11. The type I fatty acid and polyketide synthases: a tale of two megasynthases

    PubMed Central

    Tsai, Shiou-Chuan

    2008-01-01

    This review chronicles the synergistic growth of the fields of fatty acid and polyketide synthesis over the last century. In both animal fatty acid synthases and modular polyketide synthases, similar catalytic elements are covalently linked in the same order in megasynthases. Whereas in fatty acid synthases the basic elements of the design remain immutable, guaranteeing the faithful production of saturated fatty acids, in the modular polyketide synthases, the potential of the basic design has been exploited to the full for the elaboration of a wide range of secondary metabolites of extraordinary structural diversity. PMID:17898897

  12. Cryptic polyketide synthase genes in non-pathogenic Clostridium SPP.

    PubMed

    Behnken, Swantje; Hertweck, Christian

    2012-01-01

    Modular type I polyketide synthases (PKS) produce a vast array of bacterial metabolites with highly diverse biological functions. Notably, all known polyketides were isolated from aerobic bacteria, and yet no example has been reported for strict anaerobes. In this study we explored the diversity and distribution of PKS genes in the genus Clostridium. In addition to comparative genomic analyses combined with predictions of modular type I polyketide synthase (PKS) gene clusters in sequenced genomes of Clostridium spp., a representative selection of other species inhabiting a variety of ecological niches was investigated by PCR screening for PKS genes. Our data reveal that all studied pathogenic Clostridium spp. are devoid of putative PKS genes. In stark contrast, cryptic PKS genes are widespread in genomes of non-pathogenic Clostridium species. According to phylogenetic analyses, the Clostridium PKS genes have unusual and diverse origins. However, reverse transcription quantitative PCR demonstrates that these genes are silent under standard cultivation conditions, explaining why the related metabolites have been overlooked until now. This study presents clostridia as a putative source for novel bioactive polyketides.

  13. Cryptic Polyketide Synthase Genes in Non-Pathogenic Clostridium SPP

    PubMed Central

    Behnken, Swantje; Hertweck, Christian

    2012-01-01

    Modular type I polyketide synthases (PKS) produce a vast array of bacterial metabolites with highly diverse biological functions. Notably, all known polyketides were isolated from aerobic bacteria, and yet no example has been reported for strict anaerobes. In this study we explored the diversity and distribution of PKS genes in the genus Clostridium. In addition to comparative genomic analyses combined with predictions of modular type I polyketide synthase (PKS) gene clusters in sequenced genomes of Clostridium spp., a representative selection of other species inhabiting a variety of ecological niches was investigated by PCR screening for PKS genes. Our data reveal that all studied pathogenic Clostridium spp. are devoid of putative PKS genes. In stark contrast, cryptic PKS genes are widespread in genomes of non-pathogenic Clostridium species. According to phylogenetic analyses, the Clostridium PKS genes have unusual and diverse origins. However, reverse transcription quantitative PCR demonstrates that these genes are silent under standard cultivation conditions, explaining why the related metabolites have been overlooked until now. This study presents clostridia as a putative source for novel bioactive polyketides. PMID:22235310

  14. Hybrid polyketide synthases

    DOEpatents

    Fortman, Jeffrey L.; Hagen, Andrew; Katz, Leonard; Keasling, Jay D.; Poust, Sean; Zhang, Jingwei; Zotchev, Sergey

    2016-05-10

    The present invention provides for a polyketide synthase (PKS) capable of synthesizing an even-chain or odd-chain diacid or lactam or diamine. The present invention also provides for a host cell comprising the PKS and when cultured produces the even-chain diacid, odd-chain diacid, or KAPA. The present invention also provides for a host cell comprising the PKS capable of synthesizing a pimelic acid or KAPA, and when cultured produces biotin.

  15. Biosynthesis of polyketides by trans-AT polyketide synthases.

    PubMed

    Helfrich, Eric J N; Piel, Jörn

    2016-02-01

    This review discusses the biosynthesis of natural products that are generated by trans-AT polyketide synthases, a family of catalytically versatile enzymes that represents one of the major group of proteins involved in the production of bioactive polyketides. The article includes 609 references and covers the literature from 2009 through June 2015.

  16. Biosynthesis of polyketides by trans-AT polyketide synthases.

    PubMed

    Piel, Jörn

    2010-07-01

    This review discusses the biosynthesis of natural products that are generated by trans-AT polyketide synthases, a family of catalytically versatile enzymes that have recently been recognized as one of the major group of proteins involved in the production of bioactive polyketides. 436 references are cited.

  17. Evidence for an iterative module in chain elongation on the azalomycin polyketide synthase

    PubMed Central

    Hong, Hui; Sun, Yuhui; Zhou, Yongjun; Stephens, Emily; Samborskyy, Markiyan

    2016-01-01

    Summary The assembly-line synthases that produce bacterial polyketide natural products follow a modular paradigm in which each round of chain extension is catalysed by a different set or module of enzymes. Examples of deviation from this paradigm, in which a module catalyses either multiple extensions or none are of interest from both a mechanistic and an evolutionary viewpoint. We present evidence that in the biosynthesis of the 36-membered macrocyclic aminopolyol lactones (marginolactones) azalomycin and kanchanamycin, isolated respectively from Streptomyces malaysiensis DSM4137 and Streptomyces olivaceus Tü4018, the first extension module catalyses both the first and second cycles of polyketide chain extension. To confirm the integrity of the azl gene cluster, it was cloned intact on a bacterial artificial chromosome and transplanted into the heterologous host strain Streptomyces lividans, which does not possess the genes for marginolactone production. When furnished with 4-guanidinobutyramide, a specific precursor of the azalomycin starter unit, the recombinant S. lividans produced azalomycin, showing that the polyketide synthase genes in the sequenced cluster are sufficient to accomplish formation of the full-length polyketide chain. This provides strong support for module iteration in the azalomycin and kanchanamycin biosynthetic pathways. In contrast, re-sequencing of the gene cluster for biosynthesis of the polyketide β-lactone ebelactone in Streptomyces aburaviensis has shown that, contrary to a recently-published proposal, the ebelactone polyketide synthase faithfully follows the colinear modular paradigm. PMID:27829923

  18. Progress challenges and opportunities for the re-engineering of trans-AT polyketide synthases.

    PubMed

    Till, M; Race, P R

    2014-05-01

    Polyketides are a structurally and functionally diverse family of bioactive natural products that are used extensively as pharmaceuticals and agrochemicals. In bacteria these molecules are biosynthesized by giant, multi-functional enzymatic complexes, termed modular polyketide synthases (PKSs), that function in assembly-line like fashion to fuse and tailor simple carboxylic acid monomers into a vast array of elaborate chemical scaffolds. Modifying PKSs through targeted synthase re-engineering is a promising approach for accessing functionally-optimized polyketides. Due to their highly mosaic architectures the recently identified trans-AT family of modular synthases appear inherently more amenable to re-engineering than their well studied cis-AT counterparts. Here, we review recent progress in the re-engineering of trans-AT PKSs, summarize opportunities for harnessing the biosynthetic potential of these systems, and highlight challenges that such re-engineering approaches present.

  19. Structure-function analyses of plant type III polyketide synthases.

    PubMed

    Weng, Jing-Ke; Noel, Joseph P

    2012-01-01

    Plant type III polyketide synthases (PKSs) form a superfamily of biosynthetic enzymes involved in the production of a plethora of polyketide-derived natural products important for ecological adaptations and the fitness of land plants. Moreover, tremendous interest in bioengineering of type III PKSs to produce high-value compounds is increasing. Compared to type I and type II PKSs, which form either large modular protein complexes or dissociable molecular assemblies, type III PKSs exist as smaller homodimeric proteins, technically more amenable for detailed quantitative biochemical and phylogenetic analyses. In this chapter, we summarize a collection of approaches, including bioinformatics, genetics, protein crystallography, in vitro biochemistry, and mutagenesis, together affording a comprehensive interrogation of the structure-function-evolutionary relationships in the plant type III PKS family.

  20. Engineering the acyltransferase substrate specificity of assembly line polyketide synthases

    PubMed Central

    Dunn, Briana J.; Khosla, Chaitan

    2013-01-01

    Polyketide natural products act as a broad range of therapeutics, including antibiotics, immunosuppressants and anti-cancer agents. This therapeutic diversity stems from the structural diversity of these small molecules, many of which are produced in an assembly line manner by modular polyketide synthases. The acyltransferase (AT) domains of these megasynthases are responsible for selection and incorporation of simple monomeric building blocks, and are thus responsible for a large amount of the resulting polyketide structural diversity. The substrate specificity of these domains is often targeted for engineering in the generation of novel, therapeutically active natural products. This review outlines recent developments that can be used in the successful engineering of these domains, including AT sequence and structural data, mechanistic insights and the production of a diverse pool of extender units. It also provides an overview of previous AT domain engineering attempts, and concludes with proposed engineering approaches that take advantage of current knowledge. These approaches may lead to successful production of biologically active ‘unnatural’ natural products. PMID:23720536

  1. Engineering the acyltransferase substrate specificity of assembly line polyketide synthases.

    PubMed

    Dunn, Briana J; Khosla, Chaitan

    2013-08-06

    Polyketide natural products act as a broad range of therapeutics, including antibiotics, immunosuppressants and anti-cancer agents. This therapeutic diversity stems from the structural diversity of these small molecules, many of which are produced in an assembly line manner by modular polyketide synthases. The acyltransferase (AT) domains of these megasynthases are responsible for selection and incorporation of simple monomeric building blocks, and are thus responsible for a large amount of the resulting polyketide structural diversity. The substrate specificity of these domains is often targeted for engineering in the generation of novel, therapeutically active natural products. This review outlines recent developments that can be used in the successful engineering of these domains, including AT sequence and structural data, mechanistic insights and the production of a diverse pool of extender units. It also provides an overview of previous AT domain engineering attempts, and concludes with proposed engineering approaches that take advantage of current knowledge. These approaches may lead to successful production of biologically active 'unnatural' natural products.

  2. Steps towards the synthetic biology of polyketide biosynthesis

    PubMed Central

    Cummings, Matthew; Breitling, Rainer; Takano, Eriko

    2014-01-01

    Nature is providing a bountiful pool of valuable secondary metabolites, many of which possess therapeutic properties. However, the discovery of new bioactive secondary metabolites is slowing down, at a time when the rise of multidrug-resistant pathogens and the realization of acute and long-term side effects of widely used drugs lead to an urgent need for new therapeutic agents. Approaches such as synthetic biology are promising to deliver a much-needed boost to secondary metabolite drug development through plug-and-play optimized hosts and refactoring novel or cryptic bacterial gene clusters. Here, we discuss this prospect focusing on one comprehensively studied class of clinically relevant bioactive molecules, the polyketides. Extensive efforts towards optimization and derivatization of compounds via combinatorial biosynthesis and classical engineering have elucidated the modularity, flexibility and promiscuity of polyketide biosynthetic enzymes. Hence, a synthetic biology approach can build upon a solid basis of guidelines and principles, while providing a new perspective towards the discovery and generation of novel and new-to-nature compounds. We discuss the lessons learned from the classical engineering of polyketide synthases and indicate their importance when attempting to engineer biosynthetic pathways using synthetic biology approaches for the introduction of novelty and overexpression of products in a controllable manner. PMID:24372666

  3. Thioesterase Domain Swapping of a Linear Polyketide Tautomycetin with a Macrocyclic Polyketide Pikromycin in Streptomyces sp. CK4412

    PubMed Central

    Tripathi, Ashootosh; Choi, Si-Sun; Sherman, David H.; Kim, Eung-Soo

    2016-01-01

    Tautomycetin (TMC) is a linear polyketide metabolite produced by Streptomyces sp. CK4412 that has been reported to possess multiple biological functions including T cell-specific immunosuppressive and anticancer activities that occur through a mechanism of differential inhibition of protein phosphatases such as PP1, PP2A, and SHP2. We previously reported the characterization of the entire TMC biosynthetic gene cluster constituted by multifunctional type I polyketide synthase (PKS) assembly and suggested that the linear form of TMC could be generated via free acid chain termination by a narrow TMC thioesterase (TE) pocket. The modular nature of the assembly presents a unique opportunity to alter or interchange the native biosynthetic domains to produce targeted variants of TMC. Herein, we report swapping of the TMC TE domain sequence with the exact counterpart of the macrocyclic polyketide pikromycin (PIK) TE. PIK TE-swapped Streptomyces sp. CK4412 mutant produced not only TMC, but also a cyclized form of TMC, implying that the bioengineering based in vivo custom construct can be exploited to produce engineered macrolactones with new structural functionality. PMID:27277081

  4. Crystal Structures of Dehydratase Domains from the Curacin Polyketide Biosynthetic Pathway

    PubMed Central

    Akey, David L.; Razelun, Jamie R.; Tehranisa, Jason; Sherman, David H.; Gerwick, William H.; Smith, Janet L.

    2011-01-01

    SUMMARY Modular polyketide synthases (PKS) make novel natural products through a series of pre-programmed chemical steps catalyzed by an assembly line of multi-domain modules. Each assembly line step involves unique extension and modification reactions, resulting in tremendous diversity of polyketide products. Dehydratase domains catalyze formation of an α,β-double bond in the nascent polyketide intermediate. We present crystal structures of the four dehydratase domains from the curacin A PKS. The catalytic residues and substrate binding site reside in a tunnel within a single monomer. The positions of the catalytic residues and shape of the substrate tunnel explain how chirality of the substrate hydroxyl group may determine the configuration of the product double bond. Access to the active site may require opening the substrate tunnel, forming an open trench. The arrangement of monomers within the dimer is consistent among PKS dehydratases and differs from that seen in the related mammalian fatty acid synthases. PMID:20152156

  5. Architecture of the polyketide synthase module: surprises from electron cryo-microscopy

    PubMed Central

    Smith, Janet L; Skiniotis, Georgios; Sherman, David H

    2015-01-01

    Modular polyketide synthases produce a vast array of bioactive molecules that are the basis of many highly valued pharmaceuticals. The biosynthesis of these compounds is based on ordered assembly lines of multi-domain modules, each extending and modifying a specific chain-elongation intermediate before transfer to the next module for further processing. The first 3D structures of a full polyketide synthase module in different functional states were obtained recently by electron cryo-microscopy. The unexpected module architecture revealed a striking evolutionary divergence of the polyketide synthase compared to its metazoan fatty acid synthase homolog, as well as remarkable conformational rearrangements dependent on its biochemical state during the full catalytic cycle. The design and dynamics of the module are highly optimized for both catalysis and fidelity in the construction of complex, biologically active natural products. PMID:25791608

  6. Engineering of plant polyketide biosynthesis.

    PubMed

    Abe, Ikuro

    2008-11-01

    A growing number of functionally divergent the chalcone synthase (CHS) superfamily type III polyketide synthases (PKSs) have been cloned and characterized, which include recently obtained pentaketide chromone synthase (PCS) and octaketide synthase (OKS) from aloe (Aloe arborescens). Recombinant PCS expressed in Escherichia coli catalyzes iterative condensations of five molecules of malonyl-CoA to produce a pentaketide, 5,7-dihydroxy-2-methylchromone, while OKS carries out sequential condensations of eight molecules of malonyl-CoA to yield aromatic octaketides, SEK4 and SEK4b, the longest polyketides generated by the structurally simple type III PKS. The two enzymes share 91% amino acid sequence identity, maintaining most of the active-site residues of CHS including the Cys-His-Asn catalytic triad. One of the most characteristic features is that the conserved Thr197 of CHS (numbering in Medicago sativa CHS) is uniquely replaced with Met207 in PCS and with Gly207 in OKS, respectively. Site-directed mutagenesis and X-ray crystallographic studies clearly demonstrated that the chemically inert single residue lining the active-site cavity controls the polyketide chain length and the product specificity depending on the steric bulk of the side chain. Finally, on the basis of the crystal structures of both wild-type and M207G-mutant PCS, a triple mutant PCS F80A/Y82A/M207G was constructed and shown to catalyze condensations of nine molecules of malonyl-CoA to produce a novel nonaketide naphthopyrone with a fused tricyclic ring system. Structure-based engineering of the type III PKS superfamily enzymes would thus lead to further production of chemically and structurally divergent unnatural novel polyketides.

  7. Metabolic and evolutionary origin of actin-binding polyketides from diverse organisms.

    PubMed

    Ueoka, Reiko; Uria, Agustinus R; Reiter, Silke; Mori, Tetsushi; Karbaum, Petra; Peters, Eike E; Helfrich, Eric J N; Morinaka, Brandon I; Gugger, Muriel; Takeyama, Haruko; Matsunaga, Shigeki; Piel, Jörn

    2015-09-01

    Actin-targeting macrolides comprise a large, structurally diverse group of cytotoxins isolated from remarkably dissimilar micro- and macroorganisms. In spite of their disparate origins and structures, many of these compounds bind actin at the same site and exhibit structural relationships reminiscent of modular, combinatorial drug libraries. Here we investigate biosynthesis and evolution of three compound groups: misakinolides, scytophycin-type compounds and luminaolides. For misakinolides from the sponge Theonella swinhoei WA, our data suggest production by an uncultivated 'Entotheonella' symbiont, further supporting the relevance of these bacteria as sources of bioactive polyketides and peptides in sponges. Insights into misakinolide biosynthesis permitted targeted genome mining for other members, providing a cyanobacterial luminaolide producer as the first cultivated source for this dimeric compound family. The data indicate that this polyketide family is bacteria-derived and that the unusual macrolide diversity is the result of combinatorial pathway modularity for some compounds and of convergent evolution for others.

  8. Producing biofuels using polyketide synthases

    DOEpatents

    Katz, Leonard; Fortman, Jeffrey L; Keasling, Jay D

    2013-04-16

    The present invention provides for a non-naturally occurring polyketide synthase (PKS) capable of synthesizing a carboxylic acid or a lactone, and a composition such that a carboxylic acid or lactone is included. The carboxylic acid or lactone, or derivative thereof, is useful as a biofuel. The present invention also provides for a recombinant nucleic acid or vector that encodes such a PKS, and host cells which also have such a recombinant nucleic acid or vector. The present invention also provides for a method of producing such carboxylic acids or lactones using such a PKS.

  9. A Single Module Type I Polyketide Synthase Directs de Novo Macrolactone Biogenesis during Galbonolide Biosynthesis in Streptomyces galbus*

    PubMed Central

    Kim, Hyun-Ju; Karki, Suman; Kwon, So-Yeon; Park, Si-Hyung; Nahm, Baek-Hie; Kim, Yeon-Ki; Kwon, Hyung-Jin

    2014-01-01

    Galbonolide (GAL) A and B are antifungal macrolactone polyketides produced by Streptomyces galbus. During their polyketide chain assembly, GAL-A and -B incorporate methoxymalonate and methylmalonate, respectively, in the fourth chain extension step. The methoxymalonyl-acyl carrier protein biosynthesis locus (galG to K) is specifically involved in GAL-A biosynthesis, and this locus is neighbored by a gene cluster composed of galA-E. GalA-C constitute a single module, highly reducing type I polyketide synthase (PKS). GalD and GalE are cytochrome P450 and Rieske domain protein, respectively. Gene knock-out experiments verified that galB, -C, and -D are essential for GAL biosynthesis. A galD mutant accumulated a GAL-C that lacked two hydroxyl groups and a double bond when compared with GAL-B. A [U-13C]propionate feeding experiment indicated that no rare precursor other than methoxymalonate was incorporated during GAL biogenesis. A search of the S. galbus genome for a modular type I PKS system, the type that was expected to direct GAL biosynthesis, resulted in the identification of only one modular type I PKS gene cluster. Homology analysis indicated that this PKS gene cluster is the locus for vicenistatin biosynthesis. This cluster was previously reported in Streptomyces halstedii. A gene deletion of the vinP2 ortholog clearly demonstrated that this modular type I PKS system is not involved in GAL biosynthesis. Therefore, we propose that GalA-C direct macrolactone polyketide formation for GAL. Our studies provide a glimpse into a novel biochemical strategy used for polyketide synthesis; that is, the iterative assembly of propionates with highly programmed β-keto group modifications. PMID:25336658

  10. [Efficient production of polyketide products in Streptomyces hosts - A review].

    PubMed

    Yao, Yongpeng; Wang, Weishan; Yang, Keqian

    2016-03-04

    Polyketides represent an important class of structurally and functionally diverse secondary metabolites with high economic value. Among bacteria, Streptomycetes are the main producers of polyketides. To enhance polyketide production in Streptomyces hosts, rational metabolic engineering approaches have been applied, such as overexpressing rate-limiting enzymes, or transcriptional activator, increasing the supply of precursor, removing feedback inhibition by end products and heterologous expression of polyketide biosynthetic gene clusters. In this review, we discuss examples of successful metabolic engineering strategies used to improve polyketide production in Streptomycetes. Meanwhile, we also address future prospective, emerging synthetic biology strategies to dynamically adjust the metabolic fluxes of pathways related to polyketide synthesis.

  11. Screening for Expressed Nonribosomal Peptide Synthetases and Polyketide Synthases Using LC-MS/MS-Based Proteomics

    PubMed Central

    Chen, Yunqiu; McClure, Ryan A.; Kelleher, Neil L.

    2016-01-01

    Liquid chromatography–mass spectrometry (LC-MS)-based proteomics is a powerful technique for the profiling of protein expression in cells in a high-throughput fashion. Herein we report a protocol using LC-MS/MS-based proteomics for the screening of enzymes involved in natural product biosynthesis, such as nonribosomal peptide synthetases (NRPSs) and polyketide synthases (PKSs) from bacterial strains. Taking advantage of the large size of modular NRPSs and PKSs (often >200 kDa), size-based separation (SDS-PAGE) is employed prior to LC-MS/MS analysis. Based upon the protein identifications obtained through software search, we can accurately pinpoint the expressed NRPS and/or PKS gene clusters from a given strain and growth condition. The proteomics screening result can be used to guide the discovery of potentially new nonribosomal peptide and polyketide natural products. PMID:26831706

  12. Evaluating Ketoreductase Exchanges as a Means of Rationally Altering Polyketide Stereochemistry.

    PubMed

    Annaval, Thibault; Paris, Cédric; Leadlay, Peter F; Jacob, Christophe; Weissman, Kira J

    2015-06-15

    Modular polyketide synthases (PKSs) are multidomain multienzymes responsible for the biosynthesis in bacteria of a wide range of polyketide secondary metabolites of clinical value. The stereochemistry of these molecules is an attractive target for genetic engineering in attempts to produce analogues exhibiting novel therapeutic properties. The exchange of ketoreductase (KR) domains in model PKSs has been shown in several cases to predictably alter the configuration of the β-hydroxy functionalities but not of the α-methyl groups. By systematic screening of a broad panel of KR domains, we have identified two donor KRs that afford modification of α-methyl group stereochemistry. To the best of our knowledge, this provides the first direct in vivo evidence of KR-catalyzed epimerization. However, none of the introduced KRs afforded simultaneous alteration of methyl and hydroxy configurations in high yield. Therefore, swapping of whole modules might be necessary to achieve such changes in stereochemistry.

  13. Cyanobacterial polyketide synthase docking domains: a tool for engineering natural product biosynthesis.

    PubMed

    Whicher, Jonathan R; Smaga, Sarah S; Hansen, Douglas A; Brown, William C; Gerwick, William H; Sherman, David H; Smith, Janet L

    2013-11-21

    Modular type I polyketide synthases (PKSs) are versatile biosynthetic systems that initiate, successively elongate, and modify acyl chains. Intermediate transfer between modules is mediated via docking domains, which are attractive targets for PKS pathway engineering to produce natural product analogs. We identified a class 2 docking domain in cyanobacterial PKSs and determined crystal structures for two docking domain pairs, revealing a distinct class 2 docking strategy for promoting intermediate transfer. The selectivity of class 2 docking interactions, demonstrated in binding and biochemical assays, could be altered by mutagenesis. We determined the ideal fusion location for exchanging class 1 and class 2 docking domains and demonstrated effective polyketide chain transfer in heterologous modules. Thus, class 2 docking domains are tools for rational bioengineering of a broad range of PKSs containing either class 1 or 2 docking domains. Copyright © 2013 Elsevier Ltd. All rights reserved.

  14. Domain Organization and Active Site Architecture of a Polyketide Synthase C-methyltransferase.

    PubMed

    Skiba, Meredith A; Sikkema, Andrew P; Fiers, William D; Gerwick, William H; Sherman, David H; Aldrich, Courtney C; Smith, Janet L

    2016-12-16

    Polyketide metabolites produced by modular type I polyketide synthases (PKS) acquire their chemical diversity through the variety of catalytic domains within modules of the pathway. Methyltransferases are among the least characterized of the catalytic domains common to PKS systems. We determined the domain boundaries and characterized the activity of a PKS C-methyltransferase (C-MT) from the curacin A biosynthetic pathway. The C-MT catalyzes S-adenosylmethionine-dependent methyl transfer to the α-position of β-ketoacyl substrates linked to acyl carrier protein (ACP) or a small-molecule analog but does not act on β-hydroxyacyl substrates or malonyl-ACP. Key catalytic residues conserved in both bacterial and fungal PKS C-MTs were identified in a 2 Å crystal structure and validated biochemically. Analysis of the structure and the sequences bordering the C-MT provides insight into the positioning of this domain within complete PKS modules.

  15. Polyketides, toxins and pigments in Penicillium marneffei.

    PubMed

    Tam, Emily W T; Tsang, Chi-Ching; Lau, Susanna K P; Woo, Patrick C Y

    2015-10-30

    Penicillium marneffei (synonym: Talaromyces marneffei) is the most important pathogenic thermally dimorphic fungus in China and Southeastern Asia. The HIV/AIDS pandemic, particularly in China and other Southeast Asian countries, has led to the emergence of P. marneffei infection as an important AIDS-defining condition. Recently, we published the genome sequence of P. marneffei. In the P. marneffei genome, 23 polyketide synthase genes and two polyketide synthase-non-ribosomal peptide synthase hybrid genes were identified. This number is much higher than those of Coccidioides immitis and Histoplasma capsulatum, important pathogenic thermally dimorphic fungi in the Western world. Phylogenetically, these polyketide synthase genes were distributed evenly with their counterparts found in Aspergillus species and other fungi, suggesting that polyketide synthases in P. marneffei did not diverge from lineage-specific gene duplication through a recent expansion. Gene knockdown experiments and ultra-high performance liquid chromatography-photodiode array detector/electrospray ionization-quadruple time of flight-mass spectrometry analysis confirmed that at least four of the polyketide synthase genes were involved in the biosynthesis of various pigments in P. marneffei, including melanin, mitorubrinic acid, mitorubrinol, monascorubrin, rubropunctatin, citrinin and ankaflavin, some of which were mycotoxins and virulence factors of the fungus.

  16. Polyketides, Toxins and Pigments in Penicillium marneffei

    PubMed Central

    Tam, Emily W. T.; Tsang, Chi-Ching; Lau, Susanna K. P.; Woo, Patrick C. Y.

    2015-01-01

    Penicillium marneffei (synonym: Talaromyces marneffei) is the most important pathogenic thermally dimorphic fungus in China and Southeastern Asia. The HIV/AIDS pandemic, particularly in China and other Southeast Asian countries, has led to the emergence of P. marneffei infection as an important AIDS-defining condition. Recently, we published the genome sequence of P. marneffei. In the P. marneffei genome, 23 polyketide synthase genes and two polyketide synthase-non-ribosomal peptide synthase hybrid genes were identified. This number is much higher than those of Coccidioides immitis and Histoplasma capsulatum, important pathogenic thermally dimorphic fungi in the Western world. Phylogenetically, these polyketide synthase genes were distributed evenly with their counterparts found in Aspergillus species and other fungi, suggesting that polyketide synthases in P. marneffei did not diverge from lineage-specific gene duplication through a recent expansion. Gene knockdown experiments and ultra-high performance liquid chromatography-photodiode array detector/electrospray ionization-quadruple time of flight-mass spectrometry analysis confirmed that at least four of the polyketide synthase genes were involved in the biosynthesis of various pigments in P. marneffei, including melanin, mitorubrinic acid, mitorubrinol, monascorubrin, rubropunctatin, citrinin and ankaflavin, some of which were mycotoxins and virulence factors of the fungus. PMID:26529013

  17. Atlas of nonribosomal peptide and polyketide biosynthetic pathways reveals common occurrence of nonmodular enzymes.

    PubMed

    Wang, Hao; Fewer, David P; Holm, Liisa; Rouhiainen, Leo; Sivonen, Kaarina

    2014-06-24

    Nonribosomal peptides and polyketides are a diverse group of natural products with complex chemical structures and enormous pharmaceutical potential. They are synthesized on modular nonribosomal peptide synthetase (NRPS) and polyketide synthase (PKS) enzyme complexes by a conserved thiotemplate mechanism. Here, we report the widespread occurrence of NRPS and PKS genetic machinery across the three domains of life with the discovery of 3,339 gene clusters from 991 organisms, by examining a total of 2,699 genomes. These gene clusters display extraordinarily diverse organizations, and a total of 1,147 hybrid NRPS/PKS clusters were found. Surprisingly, 10% of bacterial gene clusters lacked modular organization, and instead catalytic domains were mostly encoded as separate proteins. The finding of common occurrence of nonmodular NRPS differs substantially from the current classification. Sequence analysis indicates that the evolution of NRPS machineries was driven by a combination of common descent and horizontal gene transfer. We identified related siderophore NRPS gene clusters that encoded modular and nonmodular NRPS enzymes organized in a gradient. A higher frequency of the NRPS and PKS gene clusters was detected from bacteria compared with archaea or eukarya. They commonly occurred in the phyla of Proteobacteria, Actinobacteria, Firmicutes, and Cyanobacteria in bacteria and the phylum of Ascomycota in fungi. The majority of these NRPS and PKS gene clusters have unknown end products highlighting the power of genome mining in identifying novel genetic machinery for the biosynthesis of secondary metabolites.

  18. Structural and Functional Studies of a trans-Acyltransferase Polyketide Assembly Line Enzyme that Catalyzes Stereoselective α- and β-Ketoreduction

    PubMed Central

    Piasecki, Shawn K.; Zheng, Jianting; Axelrod, Abram J.; Detelich, Madeline; Keatinge-Clay, Adrian T.

    2014-01-01

    While the cis-acyltransferase modular polyketide synthase assembly lines have largely been structurally dissected, enzymes from within the recently discovered trans-acyltransferase polyketide synthase assembly lines are just starting to be observed crystallographically. Here we examine the ketoreductase from the first polyketide synthase module of the bacillaene nonribosomal peptide synthetase/polyketide synthase at 2.35-Å resolution. This ketoreductase naturally reduces both α- and β-keto groups and is the only ketoreductase known to do so during the biosynthesis of a polyketide. The isolated ketoreductase not only reduced an N-acetylcysteamine-bound β-keto substrate to a D-β-hydroxy product, but also an N-acetylcysteamine- bound α-keto substrate to an L-α-hydroxy product. That the substrates must enter the active site from opposite directions to generate these stereochemistries suggests that the acyl-phosphopantetheine moiety is capable of accessing very different conformations despite being anchored to a serine residue of a docked acyl carrier protein. The features enabling stereocontrolled α-ketoreduction may not be extensive since a β-ketoreductase from a cis-acyltransferase polyketide synthase was identified that performs a completely stereoselective reduction of the same α-keto substrate to generate the D-α-hydroxy product. A sequence analysis of trans-acyltransferase ketoreductases reveals that a single residue, rather than a three-residue motif found in cis-acyltransferase ketoreductases, is predictive of the orientation of the resulting β-hydroxyl group. PMID:24634061

  19. Identification and characterization of the niddamycin polyketide synthase genes from Streptomyces caelestis.

    PubMed Central

    Kakavas, S J; Katz, L; Stassi, D

    1997-01-01

    The genes encoding the polyketide synthase (PKS) portion of the niddamycin biosynthetic pathway were isolated from a library of Streptomyces caelestis NRRL-2821 chromosomal DNA. Analysis of 40 kb of DNA revealed the presence of five large open reading frames (ORFs) encoding the seven modular sets of enzymatic activities required for the synthesis of a 16-membered lactone ring. The enzymatic motifs identified within each module were consistent with those predicted from the structure of niddamycin. Disruption of the second ORF of the PKS coding region eliminated niddamycin production, demonstrating that the cloned genes are involved in the biosynthesis of this compound. PMID:9393718

  20. Polyketide Double Bond Biosynthesis. Mechanistic Analysis of the Dehydratase-Containing Module 2 of the Picromycin/Methymycin Polyketide Synthase

    PubMed Central

    Wu, Jiaquan; Zaleski, Toby J.; Valenzano, Chiara; Khosla, Chaitan; Cane, David E.

    2008-01-01

    Picromycin/methymycin synthase (PICS) is a modular polyketide synthase (PKS) that is responsible for the biosynthesis of both 10-deoxymethynolide (1) and narbonolide (2), the parent 12- and 14-membered aglycone precursors of the macrolide antibiotics methymycin and picromycin, respectively. PICS module 2 is a dehydratase (DH)-containing module that catalyzes the formation of the unsaturated triketide intermediate using malonyl-CoA as the chain extension substrate. Recombinant PICS module 2+TE, with the PICS thioesterase domain appended to the C-terminus to allow release of polyketide products, was expressed in Escherichia coli. Purified PICS module 2+TE converted malonyl-CoA and 4, the N-acetylcysteamine thioester of (2S,3R)-2-methyl-3-hydroxy-pentanoic acid, to a 1:2 mixture of the triketide acid (4S,5R)-4-methyl-5-hydroxy-2-heptenoic acid (5) and (3S,4S,5R)-3,5-dihydroxy-4-methyl-n-heptanoic acid-δ-lactone (10) with a combined kcat of 0.6 min−1. The triketide lactone 10 is formed by thioesterase-catalyzed cyclization of the corresponding D-3-hydroxyacyl-SACP intermediate, a reaction which competes with dehydration catalyzed by the dehydratase domain. PICS module 2+TE showed a strong preference for the syn-diketide-SNAC 4, with a 20-fold greater kcat/Km than the anti-(2S,3S)-diketide-SNAC 14, and a 40-fold advantage over the syn-(2R,3S)-diketide-SNAC 13. PICS module 2(DH0)+TE, with an inactivated DH domain, produced exclusively 10, while three PICS module 2(KR0)+TE mutants, with inactivated KR domains, produced exclusively or predominantly the unreduced triketide ketolactone, (4S,5R)-3-oxo-4-methyl-5-hydroxy-n-heptanoic acid-δ-lactone (7). These studies establish for the first time the structure and stereochemistry of the intermediates of a polyketide chain elongation cycle catalyzed by a DH-containing module, while confirming the importance of key active site residues in both KR and DH domains. PMID:16332089

  1. A polyketide synthase-peptide synthetase gene cluster from an uncultured bacterial symbiont of Paederus beetles.

    PubMed

    Piel, Jörn

    2002-10-29

    Many drug candidates from marine and terrestrial invertebrates are suspected metabolites of uncultured bacterial symbionts. The antitumor polyketides of the pederin family, isolated from beetles and sponges, are an example. Drug development from such sources is commonly hampered by low yields and the difficulty of sustaining invertebrate cultures. To obtain insight into the true producer and find alternative supplies of these rare drug candidates, the putative pederin biosynthesis genes were cloned from total DNA of Paederus fuscipes beetles, which use this compound for chemical defense. Sequence analysis of the gene cluster and adjacent regions revealed the presence of ORFs with typical bacterial architecture and homologies. The ped cluster, which is present only in beetle specimens with high pederin content, is located on a 54-kb region bordered by transposase pseudogenes and encodes a mixed modular polyketide synthase/nonribosomal peptide synthetase. Notably, none of the modules contains regions with homology to acyltransferase domains, but two copies of isolated monodomain acyltransferase genes were found at the upstream end of the cluster. In line with an involvement in pederin biosynthesis, the upstream cluster region perfectly mirrors pederin structure. The unexpected presence of additional polyketide synthase/nonribosomal peptide synthetase modules reveals surprising insights into the evolutionary relationship between pederin-type pathways in beetles and sponges.

  2. A polyketide synthase-peptide synthetase gene cluster from an uncultured bacterial symbiont of Paederus beetles

    PubMed Central

    Piel, Jörn

    2002-01-01

    Many drug candidates from marine and terrestrial invertebrates are suspected metabolites of uncultured bacterial symbionts. The antitumor polyketides of the pederin family, isolated from beetles and sponges, are an example. Drug development from such sources is commonly hampered by low yields and the difficulty of sustaining invertebrate cultures. To obtain insight into the true producer and find alternative supplies of these rare drug candidates, the putative pederin biosynthesis genes were cloned from total DNA of Paederus fuscipes beetles, which use this compound for chemical defense. Sequence analysis of the gene cluster and adjacent regions revealed the presence of ORFs with typical bacterial architecture and homologies. The ped cluster, which is present only in beetle specimens with high pederin content, is located on a 54-kb region bordered by transposase pseudogenes and encodes a mixed modular polyketide synthase/nonribosomal peptide synthetase. Notably, none of the modules contains regions with homology to acyltransferase domains, but two copies of isolated monodomain acyltransferase genes were found at the upstream end of the cluster. In line with an involvement in pederin biosynthesis, the upstream cluster region perfectly mirrors pederin structure. The unexpected presence of additional polyketide synthase/nonribosomal peptide synthetase modules reveals surprising insights into the evolutionary relationship between pederin-type pathways in beetles and sponges. PMID:12381784

  3. Genetics of Polyketide Metabolism in Aspergillus nidulans

    PubMed Central

    Klejnstrup, Marie L.; Frandsen, Rasmus J. N.; Holm, Dorte K.; Nielsen, Morten T.; Mortensen, Uffe H.; Larsen, Thomas O.; Nielsen, Jakob B.

    2012-01-01

    Secondary metabolites are small molecules that show large structural diversity and a broad range of bioactivities. Some metabolites are attractive as drugs or pigments while others act as harmful mycotoxins. Filamentous fungi have the capacity to produce a wide array of secondary metabolites including polyketides. The majority of genes required for production of these metabolites are mostly organized in gene clusters, which often are silent or barely expressed under laboratory conditions, making discovery and analysis difficult. Fortunately, the genome sequences of several filamentous fungi are publicly available, greatly facilitating the establishment of links between genes and metabolites. This review covers the attempts being made to trigger the activation of polyketide metabolism in the fungal model organism Aspergillus nidulans. Moreover, it will provide an overview of the pathways where ten polyketide synthase genes have been coupled to polyketide products. Therefore, the proposed biosynthesis of the following metabolites will be presented; naphthopyrone, sterigmatocystin, aspyridones, emericellamides, asperthecin, asperfuranone, monodictyphenone/emodin, orsellinic acid, and the austinols. PMID:24957370

  4. Producing dicarboxylic acids using polyketide synthases

    SciTech Connect

    Katz, Leonard; Fortman, Jeffrey L; Keasling, Jay D

    2013-10-29

    The present invention provides for a polyketide synthase (PKS) capable of synthesizing a dicarboxylic acid (diacid). Such diacids include diketide-diacids and triketide-diacids. The invention includes recombinant nucleic acid encoding the PKS, and host cells comprising the PKS. The invention also includes methods for producing the diacids.

  5. Producing dicarboxylic acids using polyketide synthases

    DOEpatents

    Katz, Leonard; Fortman, Jeffrey L.; Keasling, Jay D.

    2015-05-26

    The present invention provides for a polyketide synthase (PKS) capable of synthesizing a dicarboxylic acid (diacid). Such diacids include diketide-diacids and triketide-diacids. The invention includes recombinant nucleic acid encoding the PKS, and host cells comprising the PKS. The invention also includes methods for producing the diacids.

  6. Lessons from 455 Fusarium polyketide synthases

    USDA-ARS?s Scientific Manuscript database

    In fungi, polyketide synthases (PKSs) synthesize a structurally diverse array of secondary metabolites (SMs) with a range of biological activities. The most studied SMs are toxic to animals and/or plants, alter plant growth, have beneficial pharmaceutical activities, and/or are brightly colored pigm...

  7. Mycocerosic acid synthase exemplifies the architecture of reducing polyketide synthases.

    PubMed

    Herbst, Dominik A; Jakob, Roman P; Zähringer, Franziska; Maier, Timm

    2016-03-24

    Polyketide synthases (PKSs) are biosynthetic factories that produce natural products with important biological and pharmacological activities. Their exceptional product diversity is encoded in a modular architecture. Modular PKSs (modPKSs) catalyse reactions colinear to the order of modules in an assembly line, whereas iterative PKSs (iPKSs) use a single module iteratively as exemplified by fungal iPKSs (fiPKSs). However, in some cases non-colinear iterative action is also observed for modPKSs modules and is controlled by the assembly line environment. PKSs feature a structural and functional separation into a condensing and a modifying region as observed for fatty acid synthases. Despite the outstanding relevance of PKSs, the detailed organization of PKSs with complete fully reducing modifying regions remains elusive. Here we report a hybrid crystal structure of Mycobacterium smegmatis mycocerosic acid synthase based on structures of its condensing and modifying regions. Mycocerosic acid synthase is a fully reducing iPKS, closely related to modPKSs, and the prototype of mycobacterial mycocerosic acid synthase-like PKSs. It is involved in the biosynthesis of C20-C28 branched-chain fatty acids, which are important virulence factors of mycobacteria. Our structural data reveal a dimeric linker-based organization of the modifying region and visualize dynamics and conformational coupling in PKSs. On the basis of comparative small-angle X-ray scattering, the observed modifying region architecture may be common also in modPKSs. The linker-based organization provides a rationale for the characteristic variability of PKS modules as a main contributor to product diversity. The comprehensive architectural model enables functional dissection and re-engineering of PKSs.

  8. SBSPKS: structure based sequence analysis of polyketide synthases

    PubMed Central

    Anand, Swadha; Prasad, M. V. R.; Yadav, Gitanjali; Kumar, Narendra; Shehara, Jyoti; Ansari, Md. Zeeshan; Mohanty, Debasisa

    2010-01-01

    Polyketide synthases (PKSs) catalyze biosynthesis of a diverse family of pharmaceutically important secondary metabolites. Bioinformatics analysis of sequence and structural features of PKS proteins plays a crucial role in discovery of new natural products by genome mining, as well as in design of novel secondary metabolites by biosynthetic engineering. The availability of the crystal structures of various PKS catalytic and docking domains, and mammalian fatty acid synthase module prompted us to develop SBSPKS software which consists of three major components. Model_3D_PKS can be used for modeling, visualization and analysis of 3D structure of individual PKS catalytic domains, dimeric structures for complete PKS modules and prediction of substrate specificity. Dock_Dom_Anal identifies the key interacting residue pairs in inter-subunit interfaces based on alignment of inter-polypeptide linker sequences to the docking domain structure. In case of modular PKS with multiple open reading frames (ORFs), it can predict the cognate order of substrate channeling based on combinatorial evaluation of all possible interface contacts. NRPS–PKS provides user friendly tools for identifying various catalytic domains in the sequence of a Type I PKS protein and comparing them with experimentally characterized PKS/NRPS clusters cataloged in the backend databases of SBSPKS. SBSPKS is available at http://www.nii.ac.in/sbspks.html. PMID:20444870

  9. Direct evidence that the rifamycin polyketide synthase assembles polyketide chains processively

    PubMed Central

    Yu, Tin-Wein; Shen, Yuemao; Doi-Katayama, Yukiko; Tang, Li; Park, Cheonseok; Moore, Bradley S.; Richard Hutchinson, C.; Floss, Heinz G.

    1999-01-01

    The assembly of the polyketide backbone of rifamycin B on the type I rifamycin polyketide synthase (PKS), encoded by the rifA–rifE genes, is terminated by the product of the rifF gene, an amide synthase that releases the completed undecaketide as its macrocyclic lactam. Inactivation of rifF gives a rifamycin B nonproducing mutant that still accumulates a series of linear polyketides ranging from the tetra- to a decaketide, also detected in the wild type, demonstrating that the PKS operates in a processive manner. Disruptions of the rifD module 8 and rifE module 9 and module 10 genes also result in accumulation of such linear polyketides as a consequence of premature termination of polyketide assembly. Whereas the tetraketide carries an unmodified aromatic chromophore, the penta- through decaketides have undergone oxidative cyclization to the naphthoquinone, suggesting that this modification occurs during, not after, PKS assembly. The structure of one of the accumulated compounds together with 18O experiments suggests that this oxidative cyclization produces an 8-hydroxy-7,8-dihydronaphthoquinone structure that, after the stage of proansamycin X, is dehydrogenated to an 8-hydroxynaphthoquinone. PMID:10430893

  10. Modular entanglement.

    PubMed

    Gualdi, Giulia; Giampaolo, Salvatore M; Illuminati, Fabrizio

    2011-02-04

    We introduce and discuss the concept of modular entanglement. This is the entanglement that is established between the end points of modular systems composed by sets of interacting moduli of arbitrarily fixed size. We show that end-to-end modular entanglement scales in the thermodynamic limit and rapidly saturates with the number of constituent moduli. We clarify the mechanisms underlying the onset of entanglement between distant and noninteracting quantum systems and its optimization for applications to quantum repeaters and entanglement distribution and sharing.

  11. Mechanism and engineering of polyketide chain initiation in fredericamycin biosynthesis.

    PubMed

    Das, Abhirup; Szu, Ping-Hui; Fitzgerald, Jay T; Khosla, Chaitan

    2010-07-07

    The ability to incorporate atypical primer units through the use of dedicated initiation polyketide synthase (PKS) modules offers opportunities to expand the molecular diversity of polyketide natural products. Here we identify the initiation PKS module responsible for hexadienyl priming of the antibiotic fredericamycin and investigate its biochemical properties. We also exploit this PKS module for the design and in vivo biosynthesis of unusually primed analogues of a representative polyketide product, thereby emphasizing its utility to the metabolic engineer.

  12. New insights into the formation of fungal aromatic polyketides

    PubMed Central

    Crawford, Jason M.; Townsend, Craig A.

    2011-01-01

    Fungal aromatic polyketides constitute a large family of bioactive natural products and are synthesized by the non-reducing group of iterative polyketide synthases (NR-PKSs). Their diverse structures arise from selective enzymatic modifications of reactive enzyme-bound poly-β-keto intermediates. How iterative PKSs control starter unit selection, polyketide chain initiation and elongation, intermediate folding and cyclization, selective redox or modification reactions during assembly, and product release are central mechanistic questions underlying iterative catalysis. This review highlights recent insights into these questions, with a particular focus on the biosynthetic programming of fungal aromatic polyketides, and draws comparisons to the allied biosynthetic processes in bacteria. PMID:21079635

  13. Evolution of polyketide synthesis in a Dothidiomycete forest pathogen

    USDA-ARS?s Scientific Manuscript database

    Fungal secondary metabolites have many important biological roles and some, like the toxic polyketide aflatoxin, have been intensively studied at the genetic level. Complete sets of polyketide synthase (PKS) genes can now be identified in fungal pathogens by whole genome sequencing and studied in or...

  14. PKMiner: a database for exploring type II polyketide synthases

    PubMed Central

    2012-01-01

    Background Bacterial aromatic polyketides are a pharmacologically important group of natural products synthesized by type II polyketide synthases (type II PKSs) in actinobacteria. Isolation of novel aromatic polyketides from microbial sources is currently impeded because of the lack of knowledge about prolific taxa for polyketide synthesis and the difficulties in finding and optimizing target microorganisms. Comprehensive analysis of type II PKSs and the prediction of possible polyketide chemotypes in various actinobacterial genomes will thus enable the discovery or synthesis of novel polyketides in the most plausible microorganisms. Description We performed a comprehensive computational analysis of type II PKSs and their gene clusters in actinobacterial genomes. By identifying type II PKS subclasses from the sequence analysis of 280 known type II PKSs, we developed highly accurate domain classifiers for these subclasses and derived prediction rules for aromatic polyketide chemotypes generated by different combinations of type II PKS domains. Using 319 available actinobacterial genomes, we predicted 231 type II PKSs from 40 PKS gene clusters in 25 actinobacterial genomes, and polyketide chemotypes corresponding to 22 novel PKS gene clusters in 16 genomes. These results showed that the microorganisms capable of producing aromatic polyketides are specifically distributed within a certain suborder of Actinomycetales such as Catenulisporineae, Frankineae, Micrococcineae, Micromonosporineae, Pseudonocardineae, Streptomycineae, and Streptosporangineae. Conclusions We could identify the novel candidates of type II PKS gene clusters and their polyketide chemotypes in actinobacterial genomes by comprehensive analysis of type II PKSs and prediction of aromatic polyketides. The genome analysis results indicated that the specific suborders in actinomycetes could be used as prolific taxa for polyketide synthesis. The chemotype-prediction rules with the suggested type II PKS

  15. Production of the polyketide 6-deoxyerythronolide B in the heterologous host Bacillus subtilis.

    PubMed

    Kumpfmüller, Jana; Methling, Karen; Fang, Lei; Pfeifer, Blaine A; Lalk, Michael; Schweder, Thomas

    2016-02-01

    Polyketides, such as erythromycin, are complex natural products with diverse therapeutic applications. They are synthesized by multi-modular megaenzymes, so-called polyketide synthases (PKSs). The macrolide core of erythromycin, 6-deoxyerythronolide B (6dEB), is produced by the deoxyerythronolide B synthase (DEBS) that consists of three proteins each with a size of 330-370 kDa. We cloned and investigated the expression of the corresponding gene cluster from Saccharopolyspora erythraea, which comprises more than 30 kb, in Bacillus subtilis. It is shown that the DEBS genes are functionally expressed in B. subtilis when the native eryAI-III operon was separated into three individual expression cassettes with optimized ribosomal binding sites. A synthesis of 6dEB could be detected by using the acetoin-inducible acoA promoter and a fed-batch simulating EnBase-cultivation strategy. B. subtilis was capable of the secretion of 6dEB into the medium. In order to improve the 6dEB production, several genomic modifications of this production strain were tested. This included the knockout of the native secondary metabolite clusters of B. subtilis for the synthesis of surfactin (26 kb), bacillaene (76 kb), and plipastatin (38 kb). It is revealed that the deletion of the prpBD operon, responsible for propionyl-CoA utilization, resulted in a significant increase of the 6dEB product yield when exogenous propionate is provided. Although the presented B. subtilis 6dEB production strain is not competitive with established Escherichia coli 6dEB production strains, the results of this study indicate that B. subtilis is a suitable heterologous host for the secretory production of a complex polyketide.

  16. Engineered fungal polyketide biosynthesis in Pichia pastoris: a potential excellent host for polyketide production

    PubMed Central

    2013-01-01

    Background Polyketides are one of the most important classes of secondary metabolites and usually make good drugs. Currently, heterologous production of fungal polyketides for developing a high potential industrial application system with high production capacity and pharmacutical feasibility was still at its infancy. Pichia pastoris is a highly successful system for the high production of a variety of heterologous proteins. In this work, we aim to develop a P. pastoris based in vivo fungal polyketide production system for first time and evaluate its feasibility for future industrial application. Results A recombinant P. pastoris GS115-NpgA-ATX with Aspergillus nidulans phosphopantetheinyl transferase (PPtase) gene npgA and Aspergillus terrus 6-methylsalicylic acid (6-MSA) synthase (6-MSAS) gene atX was constructed. A specific compound was isolated and idenified as 6-MSA by HPLC, LC-MS and NMR. Transcription of both genes were detected. In 5-L bioreactor, the GS115-NpgA-ATX grew well and produced 6-MSA quickly until reached a high value of 2.2 g/L by methanol induction for 20 hours. Thereafter, the cells turned to death ascribing to high concentration of antimicrobial 6-MSA. The distribution of 6-MSA changed that during early and late induction phase it existed more in supernatant while during intermediate stage it mainly located intracellular. Different from 6-MSA production strain, recombinant M. purpureus pksCT expression strains for citrinin intermediate production, no matter PksCT located in cytoplasm or in peroxisomes, did not produce any specfic compound. However, both npgA and pksCT transcripted effectively in cells and western blot analysis proved the expression of PPtase. Then the PPTase was expressed and purified, marked by fluorescent probes, and reacted with purified ACP domain and its mutant ACPm of PksCT. Fluoresence was only observed in ACP but not ACPm, indicating that the PPTase worked well with ACP to make it bioactive holo-ACP. Thus, some

  17. Narrowing the gap between the promise and reality of polyketide synthases as a synthetic biology platform

    SciTech Connect

    Poust, S; Hagen, A; Katz, L; Keasling, JD

    2014-12-01

    Engineering modular polyketide synthases (PKSs) has the potential to be an effective methodology to produce existing and novel chemicals. However, this potential has only just begun to be realized. We propose the adoption of an iterative design-build-test-learn paradigm to improve PKS engineering. We suggest methods to improve engineered PKS design by learning from laboratory-based selection; adoption of DNA design software and automation to build constructs and libraries more easily; tools for the expression of engineered proteins in a variety of heterologous hosts; and mass spectrometry-based high-throughput screening methods. Finally, lessons learned during iterations of the design-build-test-learn cycle can serve as a knowledge base for the development of a single retrosynthesis algorithm usable by both PKS experts and non-experts alike.

  18. Interrogating the Molecular Basis for Multiple Macrolactone Ring Formation by the Pikromycin Polyketide Synthase

    PubMed Central

    Kittendorf, Jeffrey D.; Beck, Brian J.; Buchholz, Tonia J.; Seufert, Wolfgang; Sherman, David H.

    2009-01-01

    Summary The pikromycin polyketide synthase (PKS) is unique in its ability to generate both 12-and 14-membered ring macrolactones. As such, dissection of the molecular basis for controlling metabolic diversity in this system remains an important objective for understanding modular PKS function and expanding chemical diversity. Here, we describe a series of experiments designed to probe the importance of the protein-protein interaction that occurs between the final two monomodules, PikAIII (module 5) and PikAIV (module 6), for the production of the 12-membered ring macrolactone 10-deoxymethynolide. The results obtained from these in vitro studies demonstrate that PikAIII and PikAIV generate the 12-membered ring macrocycle most efficiently when engaged in their native protein-protein interaction. Accordingly, the data are consistent with PikAIV adopting an alternative conformation that enables the terminal thioesterase domain to directly off-load the PikAIII-bound hexaketide intermediate for macrocyclization. PMID:17719493

  19. The biosynthesis of polyketide-derived polycyclic ethers.

    PubMed

    Gallimore, Andrew R

    2009-02-01

    The biosynthetic pathways to polyketide-derived polycyclic ethers, in bacteria, plants and marine organisms, have, until now, tended to be considered separately. The purpose of this article is to provide an integrated review of the common mechanistic aspects of polyether biosynthesis from these diverse sources. In particular, the focus will be on the proposed mechanisms of oxidative cyclisation, as well as on the known differences in polyketide chain construction between the terrestrial and marine polyethers.1 Introduction, 2 Fatty acid and polyketide biosynthesis, 3 Polyether ionophores, 4 The annonaceous acetogenins, 5 Marine polyethers, 6 Chain construction in polyether biosynthesis, 7 Acknowledgements, 8 References.

  20. A crotonyl-CoA reductase-carboxylase independent pathway for assembly of unusual alkylmalonyl-CoA polyketide synthase extender units

    PubMed Central

    Ray, Lauren; Valentic, Timothy R.; Miyazawa, Takeshi; Withall, David M.; Song, Lijiang; Milligan, Jacob C.; Osada, Hiroyuki; Takahashi, Shunji; Tsai, Shiou-Chuan; Challis, Gregory L.

    2016-01-01

    Type I modular polyketide synthases assemble diverse bioactive natural products. Such multienzymes typically use malonyl and methylmalonyl-CoA building blocks for polyketide chain assembly. However, in several cases more exotic alkylmalonyl-CoA extender units are also known to be incorporated. In all examples studied to date, such unusual extender units are biosynthesized via reductive carboxylation of α, β-unsaturated thioesters catalysed by crotonyl-CoA reductase/carboxylase (CCRC) homologues. Here we show using a chemically-synthesized deuterium-labelled mechanistic probe, and heterologous gene expression experiments that the unusual alkylmalonyl-CoA extender units incorporated into the stambomycin family of polyketide antibiotics are assembled by direct carboxylation of medium chain acyl-CoA thioesters. X-ray crystal structures of the unusual β-subunit of the acyl-CoA carboxylase (YCC) responsible for this reaction, alone and in complex with hexanoyl-CoA, reveal the molecular basis for substrate recognition, inspiring the development of methodology for polyketide bio-orthogonal tagging via incorporation of 6-azidohexanoic acid and 8-nonynoic acid into novel stambomycin analogues. PMID:28000660

  1. A crotonyl-CoA reductase-carboxylase independent pathway for assembly of unusual alkylmalonyl-CoA polyketide synthase extender units

    NASA Astrophysics Data System (ADS)

    Ray, Lauren; Valentic, Timothy R.; Miyazawa, Takeshi; Withall, David M.; Song, Lijiang; Milligan, Jacob C.; Osada, Hiroyuki; Takahashi, Shunji; Tsai, Shiou-Chuan; Challis, Gregory L.

    2016-12-01

    Type I modular polyketide synthases assemble diverse bioactive natural products. Such multienzymes typically use malonyl and methylmalonyl-CoA building blocks for polyketide chain assembly. However, in several cases more exotic alkylmalonyl-CoA extender units are also known to be incorporated. In all examples studied to date, such unusual extender units are biosynthesized via reductive carboxylation of α, β-unsaturated thioesters catalysed by crotonyl-CoA reductase/carboxylase (CCRC) homologues. Here we show using a chemically-synthesized deuterium-labelled mechanistic probe, and heterologous gene expression experiments that the unusual alkylmalonyl-CoA extender units incorporated into the stambomycin family of polyketide antibiotics are assembled by direct carboxylation of medium chain acyl-CoA thioesters. X-ray crystal structures of the unusual β-subunit of the acyl-CoA carboxylase (YCC) responsible for this reaction, alone and in complex with hexanoyl-CoA, reveal the molecular basis for substrate recognition, inspiring the development of methodology for polyketide bio-orthogonal tagging via incorporation of 6-azidohexanoic acid and 8-nonynoic acid into novel stambomycin analogues.

  2. Identification of a bioactive 51-membered macrolide complex by activation of a silent polyketide synthase in Streptomyces ambofaciens

    PubMed Central

    Laureti, Luisa; Song, Lijiang; Huang, Sheng; Corre, Christophe; Leblond, Pierre; Challis, Gregory L.; Aigle, Bertrand

    2011-01-01

    There is a constant need for new and improved drugs to combat infectious diseases, cancer, and other major life-threatening conditions. The recent development of genomics-guided approaches for novel natural product discovery has stimulated renewed interest in the search for natural product-based drugs. Genome sequence analysis of Streptomyces ambofaciens ATCC23877 has revealed numerous secondary metabolite biosynthetic gene clusters, including a giant type I modular polyketide synthase (PKS) gene cluster, which is composed of 25 genes (nine of which encode PKSs) and spans almost 150 kb, making it one of the largest polyketide biosynthetic gene clusters described to date. The metabolic product(s) of this gene cluster are unknown, and transcriptional analyses showed that it is not expressed under laboratory growth conditions. The constitutive expression of a regulatory gene within the cluster, encoding a protein that is similar to Large ATP binding of the LuxR (LAL) family proteins, triggered the expression of the biosynthetic genes. This led to the identification of four 51-membered glycosylated macrolides, named stambomycins A–D as metabolic products of the gene cluster. The structures of these compounds imply several interesting biosynthetic features, including incorporation of unusual extender units into the polyketide chain and in trans hydroxylation of the growing polyketide chain to provide the hydroxyl group for macrolide formation. Interestingly, the stambomycins possess promising antiproliferative activity against human cancer cell lines. Database searches identify genes encoding LAL regulators within numerous cryptic biosynthetic gene clusters in actinomycete genomes, suggesting that constitutive expression of such pathway-specific activators represents a powerful approach for novel bioactive natural product discovery. PMID:21444795

  3. Chapter 8. Methods for in silico prediction of microbial polyketide and nonribosomal peptide biosynthetic pathways from DNA sequence data.

    PubMed

    Bachmann, Brian O; Ravel, Jacques

    2009-01-01

    Fore-knowledge of the secondary metabolic potential of cultivated and previously uncultivated microorganisms can potentially facilitate the process of natural product discovery. By combining sequence-based knowledge with biochemical precedent, translated gene sequence data can be used to rapidly derive structural elements encoded by secondary metabolic gene clusters from microorganisms. These structural elements provide an estimate of the secondary metabolic potential of a given organism and a starting point for identification of potential lead compounds in isolation/structure elucidation campaigns. The accuracy of these predictions for a given translated gene sequence depends on the biochemistry of the metabolite class, similarity to known metabolite gene clusters, and depth of knowledge concerning its biosynthetic machinery. This chapter introduces methods for prediction of structural elements for two well-studied classes: modular polyketides and nonribosomally encoded peptides. A bioinformatics tool is presented for rapid preliminary analysis of these modular systems, and prototypical methods for converting these analyses into substructural elements are described.

  4. Pyran Rings Containing Polyketides from Penicillium raistrickii

    PubMed Central

    Ma, Li-Ying; Liu, De-Sheng; Li, De-Guo; Huang, Yu-Ling; Kang, Hui-Hui; Wang, Chun-Hua; Liu, Wei-Zhong

    2016-01-01

    Five new pyran rings containing polyketides, penicipyrans A–E (1–5), together with the known pestapyrone A (6), were isolated from the saline soil-derived Penicillium raistrickii. Their structures were determined by interpretation of NMR and HRESIMS data. The absolute configurations of compounds 4 and 5 were established by the modified Mosher’s method and single-crystal X-ray diffraction analysis, respectively. These compounds possessed high structural diversity including two α-pyrones (1, 2), three isocoumarins (3, 4, 6), and one dihydropyran derivative (5). Among them, Compound 5 exhibited cytotoxicity against HL-60 and K562 cell lines with IC50 values of 4.4 and 8.5 μM, respectively. PMID:28025533

  5. New insights into bacterial type II polyketide biosynthesis

    PubMed Central

    Zhang, Zhuan; Pan, Hai-Xue; Tang, Gong-Li

    2017-01-01

    Bacterial aromatic polyketides, exemplified by anthracyclines, angucyclines, tetracyclines, and pentangular polyphenols, are a large family of natural products with diverse structures and biological activities and are usually biosynthesized by type II polyketide synthases (PKSs). Since the starting point of biosynthesis and combinatorial biosynthesis in 1984–1985, there has been a continuous effort to investigate the biosynthetic logic of aromatic polyketides owing to the urgent need of developing promising therapeutic candidates from these compounds. Recently, significant advances in the structural and mechanistic identification of enzymes involved in aromatic polyketide biosynthesis have been made on the basis of novel genetic, biochemical, and chemical technologies. This review highlights the progress in bacterial type II PKSs in the past three years (2013–2016). Moreover, novel compounds discovered or created by genome mining and biosynthetic engineering are also included. PMID:28299197

  6. Passifloricins, polyketides alpha-pyrones from Passiflora foetida resin.

    PubMed

    Echeverri, F; Arango, V; Quiñones, W; Torres, F; Escobar, G; Rosero, Y; Archbold, R

    2001-04-01

    Three polyketides alpha-pyrones, named passifloricins, were isolated from Passiflora foetida resin; their structures and relative configurations were assigned through 2D NMR spectroscopic analyses. These types of compounds were not detected in other passion flowers.

  7. Fungal polyketide azaphilone pigments as future natural food colorants?

    PubMed

    Mapari, Sameer A S; Thrane, Ulf; Meyer, Anne S

    2010-06-01

    The recent approval of fungal carotenoids as food colorants by the European Union has strengthened the prospects for fungal cell factories for the production of polyketide pigments. Fungal production of colorants has the main advantage of making the manufacturer independent of the seasonal supply of raw materials, thus minimizing batch-to-batch variations. Here, we review the potential of polyketide pigments produced from chemotaxonomically selected non-toxigenic fungal strains (e.g. Penicillium and Epicoccum spp.) to serve as food colorants. We argue that the production of polyketide azaphilone pigments from such potentially safe hosts is advantageous over traditional processes that involve Monascus spp., which risks co-production of the mycotoxin citrinin. Thus, there is tremendous potential for the development of robust fungal production systems for polyketide pigments, both to tailor functionality and to expand the color palette of contemporary natural food colorants.

  8. Engineered biosynthesis of bacterial aromatic polyketides in Escherichia coli.

    PubMed

    Zhang, Wenjun; Li, Yanran; Tang, Yi

    2008-12-30

    Bacterial aromatic polyketides are important therapeutic compounds including front line antibiotics and anticancer drugs. It is one of the last remaining major classes of natural products of which the biosynthesis has not been reconstituted in the genetically superior host Escherichia coli. Here, we demonstrate the engineered biosynthesis of bacterial aromatic polyketides in E. coli by using a dissected and reassembled fungal polyketide synthase (PKS). The minimal PKS of the megasynthase PKS4 from Gibberella fujikuroi was extracted by using two approaches. The first approach yielded a stand-alone Ketosynthase (KS)_malonyl-CoA:ACP transferase (MAT) didomain and an acyl-carrier protein (ACP) domain, whereas the second approach yielded a compact PKS (PKS_WJ) that consists of KS, MAT, and ACP on a single polypeptide. Both minimal PKSs produced nonfungal polyketides cyclized via different regioselectivity, whereas the fungal-specific C2-C7 cyclization mode was not observed. The kinetic properties of the two minimal PKSs were characterized to confirm both PKSs can synthesize polyketides with similar efficiency as the parent PKS4 megasynthase. Both minimal PKSs interacted effectively with exogenous polyketide cyclases as demonstrated by the synthesis of predominantly PK8 3 or NonaSEK4 6 in the presence of a C9-C14 or a C7-C12 cyclase, respectively. When PKS_WJ and downstream tailoring enzymes were expressed in E. coli, the expected nonaketide anthraquinone SEK26 was recovered in good titer. High-cell density fermentation was performed to demonstrate the scale-up potential of the in vivo platform for the biosynthesis of bacterial polyketides. Using engineered fungal PKSs can therefore be a general approach toward the heterologous biosynthesis of bacterial aromatic polyketides in E. coli.

  9. Rational reprogramming of fungal polyketide first-ring cyclization

    PubMed Central

    Xu, Yuquan; Zhou, Tong; Zhou, Zhengfu; Su, Shiyou; Roberts, Sue A.; Montfort, William R.; Zeng, Jia; Chen, Ming; Zhang, Wei; Lin, Min; Zhan, Jixun; Molnár, István

    2013-01-01

    Resorcylic acid lactones and dihydroxyphenylacetic acid lactones represent important pharmacophores with heat shock response and immune system modulatory activities. The biosynthesis of these fungal polyketides involves a pair of collaborating iterative polyketide synthases (iPKSs): a highly reducing iPKS with product that is further elaborated by a nonreducing iPKS (nrPKS) to yield a 1,3-benzenediol moiety bridged by a macrolactone. Biosynthesis of unreduced polyketides requires the sequestration and programmed cyclization of highly reactive poly-β-ketoacyl intermediates to channel these uncommitted, pluripotent substrates to defined subsets of the polyketide structural space. Catalyzed by product template (PT) domains of the fungal nrPKSs and discrete aromatase/cyclase enzymes in bacteria, regiospecific first-ring aldol cyclizations result in characteristically different polyketide folding modes. However, a few fungal polyketides, including the dihydroxyphenylacetic acid lactone dehydrocurvularin, derive from a folding event that is analogous to the bacterial folding mode. The structural basis of such a drastic difference in the way a PT domain acts has not been investigated until now. We report here that the fungal vs. bacterial folding mode difference is portable on creating hybrid enzymes, and we structurally characterize the resulting unnatural products. Using structure-guided active site engineering, we unravel structural contributions to regiospecific aldol condensations and show that reshaping the cyclization chamber of a PT domain by only three selected point mutations is sufficient to reprogram the dehydrocurvularin nrPKS to produce polyketides with a fungal fold. Such rational control of first-ring cyclizations will facilitate efforts to the engineered biosynthesis of novel chemical diversity from natural unreduced polyketides. PMID:23509261

  10. Modular shield

    DOEpatents

    Snyder, Keith W.

    2002-01-01

    A modular system for containing projectiles has a sheet of material including at least a polycarbonate layer held by a metal frame having a straight frame member corresponding to each straight edge of the sheet. Each frame member has a U-shaped shield channel covering and holding a straight edge of the sheet and an adjacent U-shaped clamp channel rigidly held against the shield channel. A flexible gasket separates each sheet edge from its respective shield channel; and each frame member is fastened to each adjacent frame member only by clamps extending between adjacent clamp channels.

  11. Modular Certification

    NASA Technical Reports Server (NTRS)

    Rushby, John; Miner, Paul S. (Technical Monitor)

    2002-01-01

    Airplanes are certified as a whole: there is no established basis for separately certifying some components, particularly software-intensive ones, independently of their specific application in a given airplane. The absence of separate certification inhibits the development of modular components that could be largely "precertified" and used in several different contexts within a single airplane, or across many different airplanes. In this report, we examine the issues in modular certification of software components and propose an approach based on assume-guarantee reasoning. We extend the method from verification to certification by considering behavior in the presence of failures. This exposes the need for partitioning, and separation of assumptions and guarantees into normal and abnormal cases. We then identify three classes of property that must be verified within this framework: safe function, true guarantees, and controlled failure. We identify a particular assume-guarantee proof rule (due to McMillan) that is appropriate to the applications considered, and formally verify its soundness in PVS.

  12. Polyketide mimetics yield structural and mechanistic insights into product template domain function in nonreducing polyketide synthases.

    PubMed

    Barajas, Jesus F; Shakya, Gaurav; Moreno, Gabriel; Rivera, Heriberto; Jackson, David R; Topper, Caitlyn L; Vagstad, Anna L; La Clair, James J; Townsend, Craig A; Burkart, Michael D; Tsai, Shiou-Chuan

    2017-05-23

    Product template (PT) domains from fungal nonreducing polyketide synthases (NR-PKSs) are responsible for controlling the aldol cyclizations of poly-β-ketone intermediates assembled during the catalytic cycle. Our ability to understand the high regioselective control that PT domains exert is hindered by the inaccessibility of intrinsically unstable poly-β-ketones for in vitro studies. We describe here the crystallographic application of "atom replacement" mimetics in which isoxazole rings linked by thioethers mimic the alternating sites of carbonyls in the poly-β-ketone intermediates. We report the 1.8-Å cocrystal structure of the PksA PT domain from aflatoxin biosynthesis with a heptaketide mimetic tethered to a stably modified 4'-phosphopantetheine, which provides important empirical evidence for a previously proposed mechanism of PT-catalyzed cyclization. Key observations support the proposed deprotonation at C4 of the nascent polyketide by the catalytic His1345 and the role of a protein-coordinated water network to selectively activate the C9 carbonyl for nucleophilic addition. The importance of the 4'-phosphate at the distal end of the pantetheine arm is demonstrated to both facilitate delivery of the heptaketide mimetic deep into the PT active site and anchor one end of this linear array to precisely meter C4 into close proximity to the catalytic His1345. Additional structural features, docking simulations, and mutational experiments characterize protein-substrate mimic interactions, which likely play roles in orienting and stabilizing interactions during the native multistep catalytic cycle. These findings afford a view of a polyketide "atom-replaced" mimetic in a NR-PKS active site that could prove general for other PKS domains.

  13. Tailoring pathway modularity in the biosynthesis of erythromycin analogs heterologously engineered in E. coli.

    PubMed

    Zhang, Guojian; Li, Yi; Fang, Lei; Pfeifer, Blaine A

    2015-05-01

    Type I modular polyketide synthases are responsible for potent therapeutic compounds that include avermectin (antihelinthic), rapamycin (immunosuppressant), pikromycin (antibiotic), and erythromycin (antibiotic). However, compound access and biosynthetic manipulation are often complicated by properties of native production organisms, prompting an approach (termed heterologous biosynthesis) illustrated in this study through the reconstitution of the erythromycin pathway through Escherichia coli. Using this heterologous system, 16 tailoring pathways were introduced, systematically producing eight chiral pairs of deoxysugar substrates. Successful analog formation for each new pathway emphasizes the remarkable flexibility of downstream enzymes to accommodate molecular variation. Furthermore, analogs resulting from three of the pathways demonstrated bioactivity against an erythromycin-resistant Bacillus subtilis strain. The approach and results support a platform for continued molecular diversification of the tailoring components of this and other complex natural product pathways in a manner that mirrors the modular nature of the upstream megasynthases responsible for aglycone polyketide formation.

  14. Tailoring pathway modularity in the biosynthesis of erythromycin analogs heterologously engineered in E. coli

    PubMed Central

    Zhang, Guojian; Li, Yi; Fang, Lei; Pfeifer, Blaine A.

    2015-01-01

    Type I modular polyketide synthases are responsible for potent therapeutic compounds that include avermectin (antihelinthic), rapamycin (immunosuppressant), pikromycin (antibiotic), and erythromycin (antibiotic). However, compound access and biosynthetic manipulation are often complicated by properties of native production organisms, prompting an approach (termed heterologous biosynthesis) illustrated in this study through the reconstitution of the erythromycin pathway through Escherichia coli. Using this heterologous system, 16 tailoring pathways were introduced, systematically producing eight chiral pairs of deoxysugar substrates. Successful analog formation for each new pathway emphasizes the remarkable flexibility of downstream enzymes to accommodate molecular variation. Furthermore, analogs resulting from three of the pathways demonstrated bioactivity against an erythromycin-resistant Bacillus subtilis strain. The approach and results support a platform for continued molecular diversification of the tailoring components of this and other complex natural product pathways in a manner that mirrors the modular nature of the upstream megasynthases responsible for aglycone polyketide formation. PMID:26601183

  15. Nonribosomal peptides and polyketides of Burkholderia: new compounds potentially implicated in biocontrol and pharmaceuticals.

    PubMed

    Esmaeel, Qassim; Pupin, Maude; Jacques, Philippe; Leclère, Valérie

    2017-05-25

    Bacteria belonging to the genus Burkholderia live in various ecological niches and present a significant role in the environments through the excretion of a wide variety of secondary metabolites including modular nonribosomal peptides (NRPs) and polyketides (PKs). These metabolites represent a widely distributed biomedically and biocontrol important class of natural products including antibiotics, siderophores, and anticancers as well as biopesticides that are considered as a novel source that can be used to defend ecological niche from competitors and to promote plant growth. The aim of this review is to present all NRPs produced or potentially produced by strains of Burkholderia, as NRPs represent a major source of active compounds implicated in biocontrol. The review is a compilation of results from a large screening we have performed on 48 complete sequenced genomes available in NCBI to identify NRPS gene clusters, and data found in the literature mainly because some interesting compounds are produced by strains not yet sequenced. In addition to NRPs, hybrids NRPs/PKs are also included. Specific features about biosynthetic gene clusters and structures of the modular enzymes responsible for the synthesis, the biological activities, and the potential uses in agriculture and pharmaceutical of NRPs and hybrids NRPs/PKs will also be discussed.

  16. Synthetic biology approaches to fluorinated polyketides

    PubMed Central

    Thuronyi, Benjamin W.; Chang, Michelle C. Y.

    2016-01-01

    Conspectus The catalytic diversity of living systems offers a broad range of opportunities for developing new methods to produce small molecule targets such as fuels, materials, and pharmaceuticals. In addition to providing cost-effective and renewable methods for large-scale commercial processes, the exploration of the unusual chemical phenotypes found in living organisms can also enable the expansion of chemical space for discovery of novel function by combining orthogonal attributes from both synthetic and biological chemistry. In this context, we have focused on the development of new fluorine chemistry using synthetic biology approaches. While fluorine has become an important feature in compounds of synthetic origin, the scope of biological fluorine chemistry in living systems is limited, with fewer than 20 organofluorine natural products identified to date. In order to expand the diversity of biosynthetically accessible organofluorines, we have begun to develop methods for the site-selective introduction of fluorine into complex natural products by engineering biosynthetic machinery to incorporate fluorinated building blocks. To gain insight into how both enzyme active sites and metabolic pathways can be evolved to manage and select for fluorinated compounds, we have studied one of the only characterized natural hosts for organofluorine biosynthesis, the soil microbe Streptomyces cattleya. This information provides a template for designing engineered organofluorine enzymes, pathways, and hosts and has allowed us to initiate construction of enzymatic and cellular pathways for the production of fluorinated polyketides. PMID:25719427

  17. Type III polyketide synthases in microorganisms.

    PubMed

    Katsuyama, Yohei; Ohnishi, Yasuo

    2012-01-01

    Type III polyketide synthases (PKSs) are simple homodimers of ketosynthases which catalyze the condensation of one to several molecules of extender substrate onto a starter substrate through iterative decarboxylative Claisen condensation reactions. Type III PKSs have been found in bacteria and fungi, as well as plants. Microbial type III PKSs, which are involved in the biosynthesis of some lipidic compounds and various secondary metabolites, have several interesting characteristics that are not shared by plant type III PKSs. Further, many compounds produced by microbial type III PKSs have significant biological functions and/or important pharmaceutical activities. Thus, studies on this class of enzymes will expand our knowledge of the biosynthetic machineries that generate natural products and generate new findings about microbial physiology. The recent development of next-generation DNA sequencing has allowed for an increase in the number of microbial genomes sequenced and the discovery of many microbial type III PKS genes. Here, we describe basic methods to study microbial type III PKSs whose genes are easy to clone. Copyright © 2012 Elsevier Inc. All rights reserved.

  18. Biosynthetically Distinct Cytotoxic Polyketides from Setophoma terrestris

    PubMed Central

    El-Elimat, Tamam; Figueroa, Mario; Raja, Huzefa A.; Graf, Tyler N.; Swanson, Steven M.; Falkinham, Joseph O.; Wani, Mansukh C.; Pearce, Cedric J.

    2014-01-01

    Sixteen polyketides belonging to diverse structural classes, including monomeric/dimeric tetrahydroxanthones and resorcylic acid lactones, were isolated from an organic extract of a fungal culture Setophoma terrestris (MSX45109) using bioactivity-directed fractionation as part of a search for anticancer leads from filamentous fungi. Of these, six were new: penicillixanthone B (5), blennolide H (6), 11-deoxy blennolide D (7), blennolide I (9), blennolide J (10), and pyrenomycin (16). The known compounds were: secalonic acid A (1), secalonic acid E (2), secalonic acid G (3), penicillixanthone A (4), paecilin B (8), aigialomycin A (11), hypothemycin (12), dihydrohypothemycin (13), pyrenochaetic acid C (14), and nidulalin B (15). The structures were elucidated using a set of spectroscopic and spectrometric techniques; the absolute configurations of compounds 1–10 were determined using ECD spectroscopy combined with time-dependent density functional theory (TDDFT) calculations, while a modified Mosher’s ester method was used for compound 16. The cytotoxic activities of compounds (1–15) were evaluated using the MDA-MB-435 (melanoma) and SW-620 (colon) cancer cell lines. Compounds 1, 4, and 12 were the most potent with IC50 values ranging from 0.16 to 2.14 μM. When tested against a panel of bacteria and fungi, compounds 3 and 5 showed promising activity against the Gram-positive bacterium Micrococcus luteus with MIC values of 5 and 15 μg/mL, respectively. PMID:25574154

  19. Synthetic biology approaches to fluorinated polyketides.

    PubMed

    Thuronyi, Benjamin W; Chang, Michelle C Y

    2015-03-17

    The catalytic diversity of living systems offers a broad range of opportunities for developing new methods to produce small molecule targets such as fuels, materials, and pharmaceuticals. In addition to providing cost-effective and renewable methods for large-scale commercial processes, the exploration of the unusual chemical phenotypes found in living organisms can also enable the expansion of chemical space for discovery of novel function by combining orthogonal attributes from both synthetic and biological chemistry. In this context, we have focused on the development of new fluorine chemistry using synthetic biology approaches. While fluorine has become an important feature in compounds of synthetic origin, the scope of biological fluorine chemistry in living systems is limited, with fewer than 20 organofluorine natural products identified to date. In order to expand the diversity of biosynthetically accessible organofluorines, we have begun to develop methods for the site-selective introduction of fluorine into complex natural products by engineering biosynthetic machinery to incorporate fluorinated building blocks. To gain insight into how both enzyme active sites and metabolic pathways can be evolved to manage and select for fluorinated compounds, we have studied one of the only characterized natural hosts for organofluorine biosynthesis, the soil microbe Streptomyces cattleya. This information provides a template for designing engineered organofluorine enzymes, pathways, and hosts and has allowed us to initiate construction of enzymatic and cellular pathways for the production of fluorinated polyketides.

  20. Analysis of the cercosporin polyketide synthase CTB1 reveals a new fungal thioesterase function

    PubMed Central

    Newman, Adam G.; Vagstad, Anna L.; Belecki, Katherine; Scheerer, Jonathan R.

    2012-01-01

    The polyketide synthase CTB1 is demonstrated to catalyze pyrone formation thereby expanding the known biosynthetic repertoire of thioesterase domains in iterative, non-reducing polyketide synthases. PMID:23108075

  1. Stalk cell differentiation without polyketides in the cellular slime mold.

    PubMed

    Sato, Yukie G; Suarez, Teresa; Saito, Tamao

    2016-07-01

    Polyketides induce prestalk cell differentiation in Dictyostelium. In the double-knockout mutant of the SteelyA and B polyketide synthases, most of the pstA cells-the major part of the prestalk cells-are lost, and we show by whole mount in situ hybridization that expression of prestalk genes is also reduced. Treatment of the double-knockout mutant with the PKS inhibitor cerulenin gave a further reduction, but some pstA cells still remained in the tip region, suggesting the existence of a polyketide-independent subtype of pstA cells. The double-knockout mutant and cerulenin-treated parental Ax2 cells form fruiting bodies with fragile, single-cell layered stalks after cerulenin treatment. Our results indicate that most pstA cells are induced by polyketides, but the pstA cells at the very tip of the slug are induced in some other way. In addition, a fruiting body with a single-cell layered, vacuolated stalk can form without polyketides.

  2. Navigating the Fungal Polyketide Chemical Space: From Genes to Molecules

    PubMed Central

    Chooi, Yit-Heng; Tang, Yi

    2012-01-01

    The iterative type I polyketide synthases (IPKSs) are central to the biosynthesis of an enormously diverse array of natural products in fungi. These natural products, known as polyketides, exhibit a wide range of biological activities and include clinically important drugs as well as undesirable toxins. The PKSs synthesize these structurally diverse polyketides via a series of decarboxylative condensations of malonyl-CoA extender units and β-keto modifications in a highly programmed manner. Significant progress has been made over the past few years in understanding the biosynthetic mechanism and programming of fungal PKSs. The continuously expanding fungal genome sequence data have sparked genome-directed discoveries of new fungal PKSs and associated products. The increasing number of fungal PKSs that have been linked to their products along with in-depth biochemical and structural characterizations of these large enzymes have remarkably improved our knowledge on the molecular basis for polyketide structural diversity in fungi. This perspective highlights the recent advances and examines how the newly expanded paradigm has contributed to our ability to link fungal PKS genes to chemical structures and vice versa. The knowledge will help us navigate through the logarithmically expanding seas of genomic information for polyketide compound discovery and provided opportunities to reprogram these megasynthases to generate new chemical entities. PMID:22938194

  3. Navigating the fungal polyketide chemical space: from genes to molecules.

    PubMed

    Chooi, Yit-Heng; Tang, Yi

    2012-11-16

    The iterative type I polyketide synthases (IPKSs) are central to the biosynthesis of an enormously diverse array of natural products in fungi. These natural products, known as polyketides, exhibit a wide range of biological activities and include clinically important drugs as well as undesirable toxins. The PKSs synthesize these structurally diverse polyketides via a series of decarboxylative condensations of malonyl-CoA extender units and β-keto modifications in a highly programmed manner. Significant progress has been made over the past few years in understanding the biosynthetic mechanism and programming of fungal PKSs. The continuously expanding fungal genome sequence data have sparked genome-directed discoveries of new fungal PKSs and associated products. The increasing number of fungal PKSs that have been linked to their products along with in-depth biochemical and structural characterizations of these large enzymes have remarkably improved our knowledge on the molecular basis for polyketide structural diversity in fungi. This Perspective highlights the recent advances and examines how the newly expanded paradigm has contributed to our ability to link fungal PKS genes to chemical structures and vice versa. The knowledge will help us navigate through the logarithmically expanding seas of genomic information for polyketide compound discovery and provided opportunities to reprogram these megasynthases to generate new chemical entities.

  4. Sticky swinging arm dynamics: studies of an acyl carrier protein domain from the mycolactone polyketide synthase

    PubMed Central

    Vance, Steven; Tkachenko, Olga; Thomas, Ben; Bassuni, Mona; Hong, Hui; Nietlispach, Daniel; Broadhurst, William

    2016-01-01

    Type I modular polyketide synthases (PKSs) produce polyketide natural products by passing a growing acyl substrate chain between a series of enzyme domains housed within a gigantic multifunctional polypeptide assembly. Throughout each round of chain extension and modification reactions, the substrate stays covalently linked to an acyl carrier protein (ACP) domain. In the present study we report on the solution structure and dynamics of an ACP domain excised from MLSA2, module 9 of the PKS system that constructs the macrolactone ring of the toxin mycolactone, cause of the tropical disease Buruli ulcer. After modification of apo ACP with 4′-phosphopantetheine (Ppant) to create the holo form, 15N nuclear spin relaxation and paramagnetic relaxation enhancement (PRE) experiments suggest that the prosthetic group swings freely. The minimal chemical shift perturbations displayed by Ppant-attached C3 and C4 acyl chains imply that these substrate-mimics remain exposed to solvent at the end of a flexible Ppant arm. By contrast, hexanoyl and octanoyl chains yield much larger chemical shift perturbations, indicating that they interact with the surface of the domain. The solution structure of octanoyl-ACP shows the Ppant arm bending to allow the acyl chain to nestle into a nonpolar pocket, whereas the prosthetic group itself remains largely solvent exposed. Although the highly reduced octanoyl group is not a natural substrate for the ACP from MLSA2, similar presentation modes would permit partner enzyme domains to recognize an acyl group while it is bound to the surface of its carrier protein, allowing simultaneous interactions with both the substrate and the ACP. PMID:26920023

  5. Sticky swinging arm dynamics: studies of an acyl carrier protein domain from the mycolactone polyketide synthase.

    PubMed

    Vance, Steven; Tkachenko, Olga; Thomas, Ben; Bassuni, Mona; Hong, Hui; Nietlispach, Daniel; Broadhurst, William

    2016-04-15

    Type I modular polyketide synthases (PKSs) produce polyketide natural products by passing a growing acyl substrate chain between a series of enzyme domains housed within a gigantic multifunctional polypeptide assembly. Throughout each round of chain extension and modification reactions, the substrate stays covalently linked to an acyl carrier protein (ACP) domain. In the present study we report on the solution structure and dynamics of an ACP domain excised from MLSA2, module 9 of the PKS system that constructs the macrolactone ring of the toxin mycolactone, cause of the tropical disease Buruli ulcer. After modification of apo ACP with 4'-phosphopantetheine (Ppant) to create the holo form, (15)N nuclear spin relaxation and paramagnetic relaxation enhancement (PRE) experiments suggest that the prosthetic group swings freely. The minimal chemical shift perturbations displayed by Ppant-attached C3 and C4 acyl chains imply that these substrate-mimics remain exposed to solvent at the end of a flexible Ppant arm. By contrast, hexanoyl and octanoyl chains yield much larger chemical shift perturbations, indicating that they interact with the surface of the domain. The solution structure of octanoyl-ACP shows the Ppant arm bending to allow the acyl chain to nestle into a nonpolar pocket, whereas the prosthetic group itself remains largely solvent exposed. Although the highly reduced octanoyl group is not a natural substrate for the ACP from MLSA2, similar presentation modes would permit partner enzyme domains to recognize an acyl group while it is bound to the surface of its carrier protein, allowing simultaneous interactions with both the substrate and the ACP. © 2016 The Author(s).

  6. Modular robot

    DOEpatents

    Ferrante, Todd A.

    1997-01-01

    A modular robot may comprise a main body having a structure defined by a plurality of stackable modules. The stackable modules may comprise a manifold, a valve module, and a control module. The manifold may comprise a top surface and a bottom surface having a plurality of fluid passages contained therein, at least one of the plurality of fluid passages terminating in a valve port located on the bottom surface of the manifold. The valve module is removably connected to the manifold and selectively fluidically connects the plurality of fluid passages contained in the manifold to a supply of pressurized fluid and to a vent. The control module is removably connected to the valve module and actuates the valve module to selectively control a flow of pressurized fluid through different ones of the plurality of fluid passages in the manifold. The manifold, valve module, and control module are mounted together in a sandwich-like manner and comprise a main body. A plurality of leg assemblies are removably connected to the main body and are removably fluidically connected to the fluid passages in the manifold so that each of the leg assemblies can be selectively actuated by the flow of pressurized fluid in different ones of the plurality of fluid passages in the manifold.

  7. Modular robot

    DOEpatents

    Ferrante, T.A.

    1997-11-11

    A modular robot may comprise a main body having a structure defined by a plurality of stackable modules. The stackable modules may comprise a manifold, a valve module, and a control module. The manifold may comprise a top surface and a bottom surface having a plurality of fluid passages contained therein, at least one of the plurality of fluid passages terminating in a valve port located on the bottom surface of the manifold. The valve module is removably connected to the manifold and selectively fluidically connects the plurality of fluid passages contained in the manifold to a supply of pressurized fluid and to a vent. The control module is removably connected to the valve module and actuates the valve module to selectively control a flow of pressurized fluid through different ones of the plurality of fluid passages in the manifold. The manifold, valve module, and control module are mounted together in a sandwich-like manner and comprise a main body. A plurality of leg assemblies are removably connected to the main body and are removably fluidically connected to the fluid passages in the manifold so that each of the leg assemblies can be selectively actuated by the flow of pressurized fluid in different ones of the plurality of fluid passages in the manifold. 12 figs.

  8. Polyketide Proofreading by an Acyltransferase-like Enzyme

    PubMed Central

    Jensen, Katja; Niederkrüger, Holger; Zimmermann, Katrin; Vagstad, Anna L.; Moldenhauer, Jana; Brendel, Nicole; Frank, Sarah; Pöplau, Petra; Kohlhaas, Christoph; Townsend, Craig A.; Oldiges, Marco; Hertweck, Christian; Piel, Jörn

    2012-01-01

    SUMMARY Trans-acyltransferase polyketide synthases (trans-AT PKSs) are an important group of bacterial enzymes producing bioactive polyketides. One difference from textbook PKSs is the presence of one or more free-standing AT-like enzymes. While one homolog loads the PKS with malonyl units, the function of the second copy (AT2) was unknown. We studied the two ATs PedC and PedD involved in pederin biosynthesis in an uncultivated symbiont. PedD displayed malonyl- but not acetyltransferase activity toward various acyl carrier proteins (ACPs). In contrast, the AT2 PedC efficiently hydrolyzed acyl units bound to N-acetylcysteamine or ACP. It accepted substrates with various chain lengths and functionalizations but did not cleave malonyl-ACP. These data are consistent with the role of PedC in PKS proofreading, suggesting a similar function for other AT2 homologs and providing strategies for polyketide titer improvement and biosynthetic investigations. PMID:22444588

  9. Biodegradation of the polyketide toxin cercosporin.

    PubMed

    Mitchell, Thomas K; Chilton, William Scott; Daub, Margaret E

    2002-09-01

    Cercosporin is a non-host-specific polyketide toxin produced by many species of plant pathogens belonging to the genus Cercospora. This red-pigmented, light-activated toxin is an important pathogenicity determinant for Cercospora species. In this study, we screened 244 bacterial isolates representing 12 different genera for the ability to degrade cercosporin. Cercosporin degradation was determined by screening for the presence of cleared zones surrounding colonies on cercosporin-containing culture medium and was confirmed by assaying the kinetics of degradation in liquid medium. Bacteria belonging to four different genera exhibited the cercosporin-degrading phenotype. The isolates with the greatest cercosporin-degrading activity belonged to Xanthomonas campestris pv. zinniae and X. campestris pv. pruni. Isolates of these pathovars removed over 90% of the cercosporin from culture medium within 48 h. Bacterial degradation of red cercosporin was accompanied by a shift in the color of the growth medium to brown and then green. The disappearance of cercosporin was accompanied by the appearance of a transient green product, designated xanosporic acid. Xanosporic acid and its more stable lactone derivative, xanosporolactone, are nontoxic to cercosporin-sensitive fungi and to plant tissue and are labile in the presence of light. Detailed spectroscopic analysis (to be reported in a separate publication) of xanosporolactone revealed that cercosporin loses one methoxyl group and gains one oxygen atom in the bacterial conversion. The resulting chromophore (4,9-dihydroxy-3-oxaperlylen-10H-10-one) has never been reported before but is biosynthetically plausible via oxygen insertion by a cytochrome P-450 enzyme.

  10. Quantification of N-acetylcysteamine activated methylmalonate incorporation into polyketide biosynthesis

    PubMed Central

    Klopries, Stephan; Sundermann, Uschi

    2013-01-01

    Summary Polyketides are biosynthesized through consecutive decarboxylative Claisen condensations between a carboxylic acid and differently substituted malonic acid thioesters, both tethered to the giant polyketide synthase enzymes. Individual malonic acid derivatives are typically required to be activated as coenzyme A-thioesters prior to their enzyme-catalyzed transfer onto the polyketide synthase. Control over the selection of malonic acid building blocks promises great potential for the experimental alteration of polyketide structure and bioactivity. One requirement for this endeavor is the supplementation of the bacterial polyketide fermentation system with tailored synthetic thioester-activated malonates. The membrane permeable N-acetylcysteamine has been proposed as a coenzyme A-mimic for this purpose. Here, the incorporation efficiency into different polyketides of N-acetylcysteamine activated methylmalonate is studied and quantified, showing a surprisingly high and transferable activity of these polyketide synthase substrate analogues in vivo. PMID:23616811

  11. Evolutionary distinctiveness of fatty acid and polyketide synthesis in eukaryotes

    PubMed Central

    Kohli, Gurjeet S; John, Uwe; Van Dolah, Frances M; Murray, Shauna A

    2016-01-01

    Fatty acids, which are essential cell membrane constituents and fuel storage molecules, are thought to share a common evolutionary origin with polyketide toxins in eukaryotes. While fatty acids are primary metabolic products, polyketide toxins are secondary metabolites that are involved in ecologically relevant processes, such as chemical defence, and produce the adverse effects of harmful algal blooms. Selection pressures on such compounds may be different, resulting in differing evolutionary histories. Surprisingly, some studies of dinoflagellates have suggested that the same enzymes may catalyse these processes. Here we show the presence and evolutionary distinctiveness of genes encoding six key enzymes essential for fatty acid production in 13 eukaryotic lineages for which no previous sequence data were available (alveolates: dinoflagellates, Vitrella, Chromera; stramenopiles: bolidophytes, chrysophytes, pelagophytes, raphidophytes, dictyochophytes, pinguiophytes, xanthophytes; Rhizaria: chlorarachniophytes, haplosporida; euglenids) and 8 other lineages (apicomplexans, bacillariophytes, synurophytes, cryptophytes, haptophytes, chlorophyceans, prasinophytes, trebouxiophytes). The phylogeny of fatty acid synthase genes reflects the evolutionary history of the organism, indicating selection to maintain conserved functionality. In contrast, polyketide synthase gene families are highly expanded in dinoflagellates and haptophytes, suggesting relaxed constraints in their evolutionary history, while completely absent from some protist lineages. This demonstrates a vast potential for the production of bioactive polyketide compounds in some lineages of microbial eukaryotes, indicating that the evolution of these compounds may have played an important role in their ecological success. PMID:26784357

  12. Type III Polyketide Synthases: Discovery, Characterization, and Engineering

    ERIC Educational Resources Information Center

    Pitel, Sheryl Beth Rubin

    2009-01-01

    The polyketides are a diverse group of natural products with important applications in medicine and industry. Industry, especially the pharmaceutical industry, is under pressure to deliver "greener" chemical syntheses that are less environmentally damaging and incorporate renewable resources. There exists potential to replace current…

  13. Type III Polyketide Synthases: Discovery, Characterization, and Engineering

    ERIC Educational Resources Information Center

    Pitel, Sheryl Beth Rubin

    2009-01-01

    The polyketides are a diverse group of natural products with important applications in medicine and industry. Industry, especially the pharmaceutical industry, is under pressure to deliver "greener" chemical syntheses that are less environmentally damaging and incorporate renewable resources. There exists potential to replace current…

  14. Structural basis for biosynthetic programming of fungal aromatic polyketide cyclization.

    PubMed

    Crawford, Jason M; Korman, Tyler P; Labonte, Jason W; Vagstad, Anna L; Hill, Eric A; Kamari-Bidkorpeh, Oliver; Tsai, Shiou-Chuan; Townsend, Craig A

    2009-10-22

    Polyketides are a class of natural products with diverse structures and biological activities. The structural variability of aromatic products of fungal nonreducing, multidomain iterative polyketide synthases (NR-PKS group of IPKSs) results from regiospecific cyclizations of reactive poly-beta-keto intermediates. How poly-beta-keto species are synthesized and stabilized, how their chain lengths are determined, and, in particular, how specific cyclization patterns are controlled have been largely inaccessible and functionally unknown until recently. A product template (PT) domain is responsible for controlling specific aldol cyclization and aromatization of these mature polyketide precursors, but the mechanistic basis is unknown. Here we present the 1.8 A crystal structure and mutational studies of a dissected PT monodomain from PksA, the NR-PKS that initiates the biosynthesis of the potent hepatocarcinogen aflatoxin B(1) in Aspergillus parasiticus. Despite having minimal sequence similarity to known enzymes, the structure displays a distinct 'double hot dog' (DHD) fold. Co-crystal structures with palmitate or a bicyclic substrate mimic illustrate that PT can bind both linear and bicyclic polyketides. Docking and mutagenesis studies reveal residues important for substrate binding and catalysis, and identify a phosphopantetheine localization channel and a deep two-part interior binding pocket and reaction chamber. Sequence similarity and extensive conservation of active site residues in PT domains suggest that the mechanistic insights gleaned from these studies will prove general for this class of IPKSs, and lay a foundation for defining the molecular rules controlling NR-PKS cyclization specificity.

  15. Modular Fixturing System

    NASA Technical Reports Server (NTRS)

    Littell, Justin Anderson (Inventor); Street, Jon P. (Inventor)

    2017-01-01

    The modular fixturing system of the present invention is modular, reusable and capable of significant customization, both in terms of system radius and system height, allowing it to be arranged and rearranged in numerous unique configurations. The system includes multiple modular stanchions having stanchion shafts and stanchion feet that removably attach to apertures in a table. Angle brackets attached to the modular stanchions support shelves. These shelves in turn provide support to work pieces during fabrication processes such as welding.

  16. Portable modular detection system

    DOEpatents

    Brennan, James S.; Singh, Anup; Throckmorton, Daniel J.; Stamps, James F.

    2009-10-13

    Disclosed herein are portable and modular detection devices and systems for detecting electromagnetic radiation, such as fluorescence, from an analyte which comprises at least one optical element removably attached to at least one alignment rail. Also disclosed are modular detection devices and systems having an integrated lock-in amplifier and spatial filter and assay methods using the portable and modular detection devices.

  17. Engineered biosynthesis of plant polyketides: chain length control in an octaketide-producing plant type III polyketide synthase.

    PubMed

    Abe, Ikuro; Oguro, Satoshi; Utsumi, Yoriko; Sano, Yukie; Noguchi, Hiroshi

    2005-09-14

    The chalcone synthase (CHS) superfamily of type III polyketide synthases (PKSs) produces a variety of plant secondary metabolites with remarkable structural diversity and biological activities (e.g., chalcones, stilbenes, benzophenones, acrydones, phloroglucinols, resorcinols, pyrones, and chromones). Here we describe an octaketide-producing novel plant-specific type III PKS from aloe (Aloe arborescens) sharing 50-60% amino acid sequence identity with other plant CHS-superfamily enzymes. A recombinant enzyme expressed in Escherichia coli catalyzed seven successive decarboxylative condensations of malonyl-CoA to yield aromatic octaketides SEK4 and SEK4b, the longest polyketides known to be synthesized by the structurally simple type III PKS. Surprisingly, site-directed mutagenesis revealed that a single residue Gly207 (corresponding to the CHS's active site Thr197) determines the polyketide chain length and product specificity. Small-to-large substitutions (G207A, G207T, G207M, G207L, G207F, and G207W) resulted in loss of the octaketide-forming activity and concomitant formation of shorter chain length polyketides (from triketide to heptaketide) including a pentaketide chromone, 2,7-dihydroxy-5-methylchromone, and a hexaketide pyrone, 6-(2,4-dihydroxy-6-methylphenyl)-4-hydroxy-2-pyrone, depending on the size of the side chain. Notably, the functional diversity of the type III PKS was shown to evolve from simple steric modulation of the chemically inert single residue lining the active-site cavity accompanied by conservation of the Cys-His-Asn catalytic triad. This provided novel strategies for the engineered biosynthesis of pharmaceutically important plant polyketides.

  18. Acyl-CoA subunit selectivity in the pikromycin polyketide synthase PikAIV: steady-state kinetics and active-site occupancy analysis by FTICR-MS.

    PubMed

    Bonnett, Shilah A; Rath, Christopher M; Shareef, Abdur-Rafay; Joels, Joanna R; Chemler, Joseph A; Håkansson, Kristina; Reynolds, Kevin; Sherman, David H

    2011-09-23

    Polyketide natural products generated by type I modular polyketide synthases (PKSs) are vital components in our drug repertoire. To reprogram these biosynthetic assembly lines, we must first understand the steps that occur within the modular "black boxes." Herein, key steps of acyl-CoA extender unit selection are explored by in vitro biochemical analysis of the PikAIV PKS model system. Two complementary approaches are employed: a fluorescent-probe assay for steady-state kinetic analysis, and Fourier Transform Ion Cyclotron Resonance-mass spectrometry (FTICR-MS) to monitor active-site occupancy. Findings from five enzyme variants and four model substrates have enabled a model to be proposed involving catalysis based upon acyl-CoA substrate loading followed by differential rates of hydrolysis. These efforts suggest a strategy for future pathway engineering efforts using unnatural extender units with slow rates of hydrolytic off-loading from the acyltransferase domain. Copyright © 2011 Elsevier Ltd. All rights reserved.

  19. Conformation-Activity Relationships of Polyketide Natural Products

    PubMed Central

    Larsen, Erik M.; Wilson, Matthew R.; Taylor, Richard E.

    2015-01-01

    Polyketides represent an important class of secondary metabolites that interact with biological targets connected to a variety of disease-associated pathways. Remarkably, nature’s assembly lines, polyketide synthases, manufacture these privileged structures through a combinatorial mixture of just a few structural units. This review highlights the role of these structural elements in shaping a polyketide’s conformational preferences, the use of computer-based molecular modeling and solution NMR studies in the identification of low-energy conformers, and the importance of conformational analogues in probing the bound conformation. In particular, this review covers several examples wherein conformational analysis complements classic structure-activity relationships in the design of biologically active natural product analogues. PMID:25974024

  20. Genetic engineering of modular PKSs: from combinatorial biosynthesis to synthetic biology.

    PubMed

    Weissman, Kira J

    2016-02-01

    Multienzyme polyketide synthases (PKSs) are molecular-scale assembly lines which construct complex natural products in bacteria. The underlying modular architecture of these gigantic catalysts inspired, from the moment of their discovery, attempts to modify them by genetic engineering to produce analogues of predictable structure. These efforts have resulted in hundreds of metabolites new to nature, as detailed in this review. However, in the face of many failures, the heady days of imagining the possibilities for a truly 'combinatorial biosynthesis' of polyketides have faded. It is now more appropriate to talk about 'PKS synthetic biology' with its more modest goals of delivering specific derivatives of known structure in combination with and as a complement to synthetic chemistry approaches. The reasons for these failures will be discussed in terms of our growing understanding of the three-dimensional architectures and mechanisms of these systems. Finally, some thoughts on the future of the field will be presented.

  1. Evolutionary and functional analysis of mulberry type III polyketide synthases.

    PubMed

    Li, Han; Liang, Jiubo; Chen, Hu; Ding, Guangyu; Ma, Bi; He, Ningjia

    2016-08-04

    Type III polyketide synthases are important for the biosynthesis of flavonoids and various plant polyphenols. Mulberry plants have abundant polyphenols, but very little is known about the mulberry type III polyketide synthase genes. An analysis of these genes may provide new targets for genetic improvement to increase relevant secondary metabolites and enhance the plant tolerance to biotic and abiotic stresses. Eighteen genes encoding type III polyketide synthases were identified, including six chalcone synthases (CHS), ten stilbene synthases (STS), and two polyketide synthases (PKS). Functional characterization of four genes representing most of the MnCHS and MnSTS genes by coexpression with 4-Coumaroyl-CoA ligase in Escherichia coli indicated that their products were able to catalyze p-coumaroyl-CoA and malonyl-CoA to generate naringenin and resveratrol, respectively. Microsynteny analysis within mulberry indicated that segmental and tandem duplication events contributed to the expansion of the MnCHS family, while tandem duplications were mainly responsible for the generation of the MnSTS genes. Combining the evolution and expression analysis results of the mulberry type III PKS genes indicated that MnCHS and MnSTS genes evolved mainly under purifying selection to maintain their original functions, but transcriptional subfunctionalization occurred during long-term species evolution. Moreover, mulberry leaves can rapidly accumulated oxyresveratrol after UV-C irradiation, suggesting that resveratrol was converted to oxyresveratrol. Characterizing the functions and evolution of mulberry type III PKS genes is crucial for advancing our understanding of these genes and providing the basis for further studies on the biosynthesis of relevant secondary metabolites in mulberry plants.

  2. Phylogenetic and Structural Analysis of Polyketide Synthases in Aspergilli

    PubMed Central

    Bhetariya, Preetida J.; Prajapati, Madhvi; Bhaduri, Asani; Mandal, Rahul Shubhra; Varma, Anupam; Madan, Taruna; Singh, Yogendra; Sarma, P. Usha

    2016-01-01

    Polyketide synthases (PKSs) of Aspergillus species are multidomain and multifunctional megaenzymes that play an important role in the synthesis of diverse polyketide compounds. Putative PKS protein sequences from Aspergillus species representing medically, agriculturally, and industrially important Aspergillus species were chosen and screened for in silico studies. Six candidate Aspergillus species, Aspergillus fumigatus Af293, Aspergillus flavus NRRL3357, Aspergillus niger CBS 513.88, Aspergillus terreus NIH2624, Aspergillus oryzae RIB40, and Aspergillus clavatus NRRL1, were selected to study the PKS phylogeny. Full-length PKS proteins and only ketosynthase (KS) domain sequence were retrieved for independent phylogenetic analysis from the aforementioned species, and phylogenetic analysis was performed with characterized fungal PKS. This resulted into grouping of Aspergilli PKSs into nonreducing (NR), partially reducing (PR), and highly reducing (HR) PKS enzymes. Eight distinct clades with unique domain arrangements were classified based on homology with functionally characterized PKS enzymes. Conserved motif signatures corresponding to each type of PKS were observed. Three proteins from Protein Data Bank corresponding to NR, PR, and HR type of PKS (XP_002384329.1, XP_753141.2, and XP_001402408.2, respectively) were selected for mapping of conserved motifs on three-dimensional structures of KS domain. Structural variations were found at the active sites on modeled NR, PR, and HR enzymes of Aspergillus. It was observed that the number of iteration cycles was dependent on the size of the cavity in the active site of the PKS enzyme correlating with a type with reducing or NR products, such as pigment, 6MSA, and lovastatin. The current study reports the grouping and classification of PKS proteins of Aspergilli for possible exploration of novel polyketides based on sequence homology; this information can be useful for selection of PKS for polyketide exploration and

  3. Allergens/Antigens, toxins and polyketides of important Aspergillus species.

    PubMed

    Bhetariya, Preetida J; Madan, Taruna; Basir, Seemi Farhat; Varma, Anupam; Usha, Sarma P

    2011-04-01

    The medical, agricultural and biotechnological importance of the primitive eukaryotic microorganisms, the Fungi was recognized way back in 1920. Among various groups of fungi, the Aspergillus species are studied in great detail using advances in genomics and proteomics to unravel biological and molecular mechanisms in these fungi. Aspergillus fumigatus, Aspergillus flavus, Aspergillus niger, Aspergillus parasiticus, Aspergillus nidulans and Aspergillus terreus are some of the important species relevant to human, agricultural and biotechnological applications. The potential of Aspergillus species to produce highly diversified complex biomolecules such as multifunctional proteins (allergens, antigens, enzymes) and polyketides is fascinating and demands greater insight into the understanding of these fungal species for application to human health. Recently a regulator gene for secondary metabolites, LaeA has been identified. Gene mining based on LaeA has facilitated new metabolites with antimicrobial activity such as emericellamides and antitumor activity such as terrequinone A from A. nidulans. Immunoproteomic approach was reported for identification of few novel allergens for A. fumigatus. In this context, the review is focused on recent developments in allergens, antigens, structural and functional diversity of the polyketide synthases that produce polyketides of pharmaceutical and biological importance. Possible antifungal drug targets for development of effective antifungal drugs and new strategies for development of molecular diagnostics are considered.

  4. Polyketide proofreading by an acyltransferase-like enzyme.

    PubMed

    Jensen, Katja; Niederkrüger, Holger; Zimmermann, Katrin; Vagstad, Anna L; Moldenhauer, Jana; Brendel, Nicole; Frank, Sarah; Pöplau, Petra; Kohlhaas, Christoph; Townsend, Craig A; Oldiges, Marco; Hertweck, Christian; Piel, Jörn

    2012-03-23

    Trans-acyltransferase polyketide synthases (trans-AT PKSs) are an important group of bacterial enzymes producing bioactive polyketides. One difference from textbook PKSs is the presence of one or more free-standing AT-like enzymes. While one homolog loads the PKS with malonyl units, the function of the second copy (AT2) was unknown. We studied the two ATs PedC and PedD involved in pederin biosynthesis in an uncultivated symbiont. PedD displayed malonyl- but not acetyltransferase activity toward various acyl carrier proteins (ACPs). In contrast, the AT2 PedC efficiently hydrolyzed acyl units bound to N-acetylcysteamine or ACP. It accepted substrates with various chain lengths and functionalizations but did not cleave malonyl-ACP. These data are consistent with the role of PedC in PKS proofreading, suggesting a similar function for other AT2 homologs and providing strategies for polyketide titer improvement and biosynthetic investigations.

  5. Phylogenomic analysis of polyketide synthase genes in actinomycetes: structural analysis of KS domains and modules of polyketide synthases.

    PubMed

    Sarwar, Samreen; Ahmed, Mehboob; Hasnain, Shahida

    2012-01-01

    Polyketides are complex and diverse secondary metabolites, synthesised by large multifunctional enzymes, Polyketide Synthases (PKS). The phylogenomic analysis of β-ketosynthase (KS) domains and PKSs within actinomycetes suggests the contribution of point mutations, gene duplications, horizontal gene transfer and homologous recombination in the evolution of PKSs. PKS genealogy suggested the ancestral module structure with KS-AT-ACP domain composition. KS domains showed similar core and highly variable loop regions at the dimer interface, which seems to affect the selectivity of the primer unit. In PKS modules, the linker regions comprise a significant fraction of the module. The reducing domains (ketoreductase and dehydrogenase) protrude out from the central axis of the module and also responsible for extreme variability in the final products. Thus, phylogenomic and structural analysis of PKSs can assist in the artificial reprogramming of PKSs.

  6. Second-generation probes for biosynthetic intermediate capture: towards a comprehensive profiling of polyketide assembly† †Electronic supplementary information (ESI) available: General methods for the synthesis of chemical probes and LC-HRMS analysis of the biosynthetic intermediates isolated from S. lasaliensis. See DOI: 10.1039/c6cc04681a Click here for additional data file.

    PubMed Central

    Wilkening, Ina; Gazzola, Silvia; Riva, Elena; Parascandolo, James S.; Song, Lijiang

    2016-01-01

    Malonyl carba(dethia) N-decanoyl cysteamine methyl esters and novel acetoxymethyl esters were utilised as second-generation probes for polyketide intermediate capture. The use of these tools in vivo led to the characterisation of an almost complete set of biosynthetic intermediates from a modular assembly line, providing a first kinetic overview of intermediate processing leading to complex natural product formation. PMID:27481638

  7. Development of an analysis program of type I polyketide synthase gene clusters using homology search and profile hidden Markov model.

    PubMed

    Tae, Hongseok; Sohng, Jae Kyung; Park, Kiejung

    2009-02-01

    MAPSI (Management and Analysis for Polyketide Synthase Type I) has been developed to offer computational analysis methods to detect type I PKS (polyketide synthase) gene clusters in genome sequences. MAPSI provides a genome analysis component, which detects PKS gene clusters by identifying domains in proteins of a genome. MAPSI also contains databases on polyketides and genome annotation data, as well as analytic components such as new PKS assembly and domain analysis. The polyketide data and analysis component are accessible through Web interfaces and are displayed with diverse information. MAPSI, which was developed to aid researchers studying type I polyketides, provides diverse components to access and analyze polyketide information and should become a very powerful computational tool for polyketide research. The system can be extended through further studies of factors related to the biological activities of polyketides.

  8. Synthetic strategy of nonreducing iterative polyketide synthases and the origin of the classical starter-unit effect

    USDA-ARS?s Scientific Manuscript database

    Polyketide natural products are found in bacteria, fungi, and plants. They contribute disproportionately to the arsenal of clinically used agents, stimulating investigation of their biosynthetic pathways for purposes of improved production and metabolic engineering. Polyketides isolated from fungi...

  9. Implementing Modular A Levels.

    ERIC Educational Resources Information Center

    Holding, Gordon

    This document, which is designed for curriculum managers at British further education (FE) colleges, presents basic information on the implementation and perceived benefits of the General Certificate of Education (GCE) modular A (Advanced) levels. The information was synthesized from a survey of 12 FE colleges that introduced the modular A levels…

  10. Modular tokamak configuration

    SciTech Connect

    Thomson, S.L.

    1985-01-01

    This report is concerned with the modular tokamak configuration, and presents information on the following topics: modularity; external vacuum boundary; vertical maintenance; combined reactor building/biological shield with totally remote maintenance; independent TF coils; minimum TF coil bore; saddle PF coils; and heat transport system in bore.

  11. Modular Buildings Buying Guide.

    ERIC Educational Resources Information Center

    Morris, Susan

    1991-01-01

    Suggests that child care program directors who are expanding their programs or opening new child care centers investigate the possibility of renting, leasing, or purchasing a modular building. Discusses the advantages of modular buildings over conventional building construction or rented space in an occupied building. Provides information about…

  12. Algal carbohydrates affect polyketide synthesis of the lichen-forming fungus Cladonia rangiferina.

    PubMed

    Elshobary, Mostafa E; Osman, Mohamed E; Abo-Shady, Atef M; Komatsu, Emy; Perreault, Hélène; Sorensen, John; Piercey-Normore, Michele D

    2016-01-01

    Lichen secondary metabolites (polyketides) are produced by the fungal partner, but the role of algal carbohydrates in polyketide biosynthesis is not clear. This study examined whether the type and concentration of algal carbohydrate explained differences in polyketide production and gene transcription by a lichen fungus (Cladonia rangiferina). The carbohydrates identified from a free-living cyanobacterium (Spirulina platensis; glucose), a lichen-forming alga (Diplosphaera chodatii; sorbitol) and the lichen alga that associates with C. rangiferina (Asterochloris sp.; ribitol) were used in each of 1%, 5% and 10% concentrations to enrich malt yeast extract media for culturing the mycobiont. Polyketides were determined by high performance liquid chromatography (HPLC), and polyketide synthase (PKS) gene transcription was measured by quantitative PCR of the ketosynthase domain of four PKS genes. The lower concentrations of carbohydrates induced the PKS gene expression where ribitol up-regulated CrPKS1 and CrPKS16 gene transcription and sorbitol up-regulated CrPKS3 and CrPKS7 gene transcription. The HPLC results revealed that lower concentrations of carbon sources increased polyketide production for three carbohydrates. One polyketide from the natural lichen thallus (fumarprotocetraric acid) also was produced by the fungal culture in ribitol supplemented media only. This study provides a better understanding of the role of the type and concentration of the carbon source in fungal polyketide biosynthesis in the lichen Cladonia rangiferina. © 2016 by The Mycological Society of America.

  13. A diverse family of Type III polyketide synthases in Eucalyptus species.

    PubMed

    Rubin-Pitel, Sheryl B; Luo, Yunzi; Lee, Jung-Kul; Zhao, Huimin

    2010-08-01

    Eucalyptus species synthesize a wealth of polyketide natural products, but no relevant biosynthetic enzyme has been identified. Degenerate primers designed from conserved regions of fourteen chalcone synthase superfamily enzymes were used to isolate gene fragments from at least five different Type III polyketide synthases (PKSs) in E. camaldulensis and E. robusta.

  14. A new polyketide, penicillolide from the marine-derived fungus Penicillium sacculum.

    PubMed

    Liu, Tao; Zhang, Songya; Li, Zhanlin; Wang, Yu; Chen, Zaixing; Bai, Jiao; Tian, Li; Pei, Yuehu; Hua, Huiming

    2016-01-01

    A new polyketide, penicillolide (1) was isolated from the fermentation broth of the marine-derived fungus Penicillium sacculum GT-308. Compound 1 is a polyketide with a unique carbon skeleton. The structure of this compound was established via extensive spectroscopic analyses including 1D-, 2D-NMR, and HRESI-MS.

  15. Bioinformatics Prediction of Polyketide Synthase Gene Clusters from Mycosphaerella fijiensis

    PubMed Central

    Noar, Roslyn D.; Daub, Margaret E.

    2016-01-01

    Mycosphaerella fijiensis, causal agent of black Sigatoka disease of banana, is a Dothideomycete fungus closely related to fungi that produce polyketides important for plant pathogenicity. We utilized the M. fijiensis genome sequence to predict PKS genes and their gene clusters and make bioinformatics predictions about the types of compounds produced by these clusters. Eight PKS gene clusters were identified in the M. fijiensis genome, placing M. fijiensis into the 23rd percentile for the number of PKS genes compared to other Dothideomycetes. Analysis of the PKS domains identified three of the PKS enzymes as non-reducing and two as highly reducing. Gene clusters contained types of genes frequently found in PKS clusters including genes encoding transporters, oxidoreductases, methyltransferases, and non-ribosomal peptide synthases. Phylogenetic analysis identified a putative PKS cluster encoding melanin biosynthesis. None of the other clusters were closely aligned with genes encoding known polyketides, however three of the PKS genes fell into clades with clusters encoding alternapyrone, fumonisin, and solanapyrone produced by Alternaria and Fusarium species. A search for homologs among available genomic sequences from 103 Dothideomycetes identified close homologs (>80% similarity) for six of the PKS sequences. One of the PKS sequences was not similar (< 60% similarity) to sequences in any of the 103 genomes, suggesting that it encodes a unique compound. Comparison of the M. fijiensis PKS sequences with those of two other banana pathogens, M. musicola and M. eumusae, showed that these two species have close homologs to five of the M. fijiensis PKS sequences, but three others were not found in either species. RT-PCR and RNA-Seq analysis showed that the melanin PKS cluster was down-regulated in infected banana as compared to growth in culture. Three other clusters, however were strongly upregulated during disease development in banana, suggesting that they may encode

  16. Bioinformatics Prediction of Polyketide Synthase Gene Clusters from Mycosphaerella fijiensis.

    PubMed

    Noar, Roslyn D; Daub, Margaret E

    2016-01-01

    Mycosphaerella fijiensis, causal agent of black Sigatoka disease of banana, is a Dothideomycete fungus closely related to fungi that produce polyketides important for plant pathogenicity. We utilized the M. fijiensis genome sequence to predict PKS genes and their gene clusters and make bioinformatics predictions about the types of compounds produced by these clusters. Eight PKS gene clusters were identified in the M. fijiensis genome, placing M. fijiensis into the 23rd percentile for the number of PKS genes compared to other Dothideomycetes. Analysis of the PKS domains identified three of the PKS enzymes as non-reducing and two as highly reducing. Gene clusters contained types of genes frequently found in PKS clusters including genes encoding transporters, oxidoreductases, methyltransferases, and non-ribosomal peptide synthases. Phylogenetic analysis identified a putative PKS cluster encoding melanin biosynthesis. None of the other clusters were closely aligned with genes encoding known polyketides, however three of the PKS genes fell into clades with clusters encoding alternapyrone, fumonisin, and solanapyrone produced by Alternaria and Fusarium species. A search for homologs among available genomic sequences from 103 Dothideomycetes identified close homologs (>80% similarity) for six of the PKS sequences. One of the PKS sequences was not similar (< 60% similarity) to sequences in any of the 103 genomes, suggesting that it encodes a unique compound. Comparison of the M. fijiensis PKS sequences with those of two other banana pathogens, M. musicola and M. eumusae, showed that these two species have close homologs to five of the M. fijiensis PKS sequences, but three others were not found in either species. RT-PCR and RNA-Seq analysis showed that the melanin PKS cluster was down-regulated in infected banana as compared to growth in culture. Three other clusters, however were strongly upregulated during disease development in banana, suggesting that they may encode

  17. Assembly Line Polyketide Synthases: Mechanistic Insights and Unsolved Problems

    PubMed Central

    2015-01-01

    Two hallmarks of assembly line polyketide synthases have motivated an interest in these unusual multienzyme systems, their stereospecificity and their capacity for directional biosynthesis. In this review, we summarize the state of knowledge regarding the mechanistic origins of these two remarkable features, using the 6-deoxyerythronolide B synthase as a prototype. Of the 10 stereocenters in 6-deoxyerythronolide B, the stereochemistry of nine carbon atoms is directly set by ketoreductase domains, which catalyze epimerization and/or diastereospecific reduction reactions. The 10th stereocenter is established by the sequential action of three enzymatic domains. Thus, the problem has been reduced to a challenge in mainstream enzymology, where fundamental gaps remain in our understanding of the structural basis for this exquisite stereochemical control by relatively well-defined active sites. In contrast, testable mechanistic hypotheses for the phenomenon of vectorial biosynthesis are only just beginning to emerge. Starting from an elegant theoretical framework for understanding coupled vectorial processes in biology [Jencks, W. P. (1980) Adv. Enzymol. Relat. Areas Mol. Biol. 51, 75–106], we present a simple model that can explain assembly line polyketide biosynthesis as a coupled vectorial process. Our model, which highlights the important role of domain–domain interactions, not only is consistent with recent observations but also is amenable to further experimental verification and refinement. Ultimately, a definitive view of the coordinated motions within and between polyketide synthase modules will require a combination of structural, kinetic, spectroscopic, and computational tools and could be one of the most exciting frontiers in 21st Century enzymology. PMID:24779441

  18. Assembly line polyketide synthases: mechanistic insights and unsolved problems.

    PubMed

    Khosla, Chaitan; Herschlag, Daniel; Cane, David E; Walsh, Christopher T

    2014-05-13

    Two hallmarks of assembly line polyketide synthases have motivated an interest in these unusual multienzyme systems, their stereospecificity and their capacity for directional biosynthesis. In this review, we summarize the state of knowledge regarding the mechanistic origins of these two remarkable features, using the 6-deoxyerythronolide B synthase as a prototype. Of the 10 stereocenters in 6-deoxyerythronolide B, the stereochemistry of nine carbon atoms is directly set by ketoreductase domains, which catalyze epimerization and/or diastereospecific reduction reactions. The 10th stereocenter is established by the sequential action of three enzymatic domains. Thus, the problem has been reduced to a challenge in mainstream enzymology, where fundamental gaps remain in our understanding of the structural basis for this exquisite stereochemical control by relatively well-defined active sites. In contrast, testable mechanistic hypotheses for the phenomenon of vectorial biosynthesis are only just beginning to emerge. Starting from an elegant theoretical framework for understanding coupled vectorial processes in biology [Jencks, W. P. (1980) Adv. Enzymol. Relat. Areas Mol. Biol. 51, 75-106], we present a simple model that can explain assembly line polyketide biosynthesis as a coupled vectorial process. Our model, which highlights the important role of domain-domain interactions, not only is consistent with recent observations but also is amenable to further experimental verification and refinement. Ultimately, a definitive view of the coordinated motions within and between polyketide synthase modules will require a combination of structural, kinetic, spectroscopic, and computational tools and could be one of the most exciting frontiers in 21st Century enzymology.

  19. Biosynthesis of oxygen and nitrogen-containing heterocycles in polyketides.

    PubMed

    Hemmerling, Franziska; Hahn, Frank

    2016-01-01

    This review highlights the biosynthesis of heterocycles in polyketide natural products with a focus on oxygen and nitrogen-containing heterocycles with ring sizes between 3 and 6 atoms. Heterocycles are abundant structural elements of natural products from all classes and they often contribute significantly to their biological activity. Progress in recent years has led to a much better understanding of their biosynthesis. In this context, plenty of novel enzymology has been discovered, suggesting that these pathways are an attractive target for future studies.

  20. Biosynthesis of oxygen and nitrogen-containing heterocycles in polyketides

    PubMed Central

    Hemmerling, Franziska

    2016-01-01

    Summary This review highlights the biosynthesis of heterocycles in polyketide natural products with a focus on oxygen and nitrogen-containing heterocycles with ring sizes between 3 and 6 atoms. Heterocycles are abundant structural elements of natural products from all classes and they often contribute significantly to their biological activity. Progress in recent years has led to a much better understanding of their biosynthesis. In this context, plenty of novel enzymology has been discovered, suggesting that these pathways are an attractive target for future studies. PMID:27559404

  1. Engineered biosynthesis of plant polyketides: manipulation of chalcone synthase.

    PubMed

    Abe, Ikuro; Watanabe, Tatsuya; Morita, Hiroyuki; Kohno, Toshiyuki; Noguchi, Hiroshi

    2006-02-02

    [reaction: see text]. Chalcone synthase (CHS) is a plant-specific type III polyketide synthase catalyzing condensation of 4-coumaroyl-CoA with three molecules of malonyl-CoA. Surprisingly, it was demonstrated that S338V mutant of Scutellaria baicalensis CHS produced octaketides SEK4/SEK4b from eight molecules of malonyl-CoA. Further, the octaketides-forming activity was dramatically increased in a CHS triple mutant (T197G/G256L/S338T). The functional conversion is based on the simple steric modulation of a chemically inert residue lining the active-site cavity.

  2. Producing a trimethylpentanoic acid using hybrid polyketide synthases

    DOEpatents

    Katz, Leonard; Fortman, Jeffrey L; Keasling, Jay D

    2014-10-07

    The present invention provides for a polyketide synthase (PKS) capable of synthesizing trimethylpentanoic acid. The present invention also provides for a host cell comprising the PKS and when cultured produces the trimethylpentanoic acid. The present invention also provides for a method of producing the trimethylpentanoic acid, comprising: providing a host cell of the present invention, and culturing said host cell in a suitable culture medium such that the trimethylpentanoic acid is produced, optionally isolating the trimethylpentanoic acid, and optionally, reducing the isolated trimethylpentanoic acid into a trimethylpentanol or an iso-octane.

  3. Modular kinetic analysis.

    PubMed

    Krab, Klaas

    2011-01-01

    Modularization is an important strategy to tackle the study of complex biological systems. Modular kinetic analysis (MKA) is a quantitative method to extract kinetic information from such a modularized system that can be used to determine the control and regulatory structure of the system, and to pinpoint and quantify the interaction of effectors with the system. The principles of the method are described, and the relation with metabolic control analysis is discussed. Examples of application of MKA are given. Copyright © 2011 Elsevier Inc. All rights reserved.

  4. Insights from the Sea: Structural Biology of Marine Polyketide Synthases

    PubMed Central

    Akey, David L.; Gehret, Jennifer J.; Khare, Dheeraj; Smith, Janet L.

    2013-01-01

    The world’s oceans are a rich source of natural products with extremely interesting chemistry. Biosynthetic pathways have been worked out for a few, and the story is being enriched with crystal structures of interesting pathway enzymes. By far, the greatest number of structural insights from marine biosynthetic pathways has originated with studies of curacin A, a poster child for interesting marine chemistry with its cyclopropane and thiazoline rings, internal cis double bond, and terminal alkene. Using the curacin A pathway as a model, structural details are now available for a novel loading enzyme with remarkable dual decarboxylase and acetyltransferase activities, an Fe2+/α-ketoglutarate-dependent halogenase that dictates substrate binding order through conformational changes, a decarboxylase that establishes regiochemistry for cyclopropane formation, and a thioesterase with specificity for β-sulfated substrates that lead to terminal alkene offloading. The four curacin A pathway dehydratases reveal an intrinsic flexibility that may accommodate bulky or stiff polyketide intermediates. In the salinosporamide A pathway, active site volume determines the halide specificity of a halogenase that catalyzes for the synthesis of a halogenated building block. Structures of a number of putative polyketide cyclases may help in understanding reaction mechanisms and substrate specificities although their substrates are presently unknown. PMID:22498975

  5. Insights from the sea: structural biology of marine polyketide synthases.

    PubMed

    Akey, David L; Gehret, Jennifer J; Khare, Dheeraj; Smith, Janet L

    2012-10-01

    The world's oceans are a rich source of natural products with extremely interesting chemistry. Biosynthetic pathways have been worked out for a few, and the story is being enriched with crystal structures of interesting pathway enzymes. By far, the greatest number of structural insights from marine biosynthetic pathways has originated with studies of curacin A, a poster child for interesting marine chemistry with its cyclopropane and thiazoline rings, internal cis double bond, and terminal alkene. Using the curacin A pathway as a model, structural details are now available for a novel loading enzyme with remarkable dual decarboxylase and acetyltransferase activities, an Fe(2+)/α-ketoglutarate-dependent halogenase that dictates substrate binding order through conformational changes, a decarboxylase that establishes regiochemistry for cyclopropane formation, and a thioesterase with specificity for β-sulfated substrates that lead to terminal alkene offloading. The four curacin A pathway dehydratases reveal an intrinsic flexibility that may accommodate bulky or stiff polyketide intermediates. In the salinosporamide A pathway, active site volume determines the halide specificity of a halogenase that catalyzes for the synthesis of a halogenated building block. Structures of a number of putative polyketide cyclases may help in understanding reaction mechanisms and substrate specificities although their substrates are presently unknown.

  6. Zosteropenillines: Polyketides from the Marine-Derived Fungus Penicillium thomii

    PubMed Central

    Afiyatullov, Shamil Sh.; Leshchenko, Elena V.; Berdyshev, Dmitrii V.; Sobolevskaya, Maria P.; Antonov, Alexandr S.; Denisenko, Vladimir A.; Popov, Roman S.; Pivkin, Mikhail V.; Udovenko, Anatoly A.; Pislyagin, Evgeny A.; von Amsberg, Gunhild; Dyshlovoy, Sergey A.

    2017-01-01

    Twelve new polyketides, zosteropenillines A–L (1–12), together with known polyketide pallidopenilline A (13), were isolated from the ethylacetate extract of the fungus Penicillium thomii associated with the seagrass Zostera marina. Their structures were established based on spectroscopic methods. The absolute configuration of zosteropenilline A (1) as 4R, 5S, 8S, 9R, 10R, and 13S was determined by a combination of the modified Mosher’s method, X-ray analysis, and NOESY data. Absolute configurations of zosteropenillines B–D (2–4) were determined by time-dependent density functional theory (TD-DFT) calculations of ECD spectra. The effect of compounds 1–3, 7, 8, 10, and 11 on the viability of human drug-resistant prostate cancer cells PC3 as well as on autophagy in these cancer cells and inhibitory effects of compounds 1, 2, and 8–10 on NO production in LPS-induced RAW 264.7 murine macrophages were examined. PMID:28218691

  7. Modular tokamak magnetic system

    DOEpatents

    Yang, Tien-Fang

    1988-01-01

    A modular tokamak system comprised of a plurality of interlocking moldules. Each module is comprised of a vacuum vessel section, a toroidal field coil, moldular saddle coils which generate a poloidal magnetic field and ohmic heating coils.

  8. Successful modular cosmology

    NASA Astrophysics Data System (ADS)

    Kadota, Kenji; Stewart, Ewan D.

    2003-07-01

    We present a modular cosmology scenario where the difficulties encountered in conventional modular cosmology are solved in a self-consistent manner, with definite predictions to be tested by observation. Notably, the difficulty of the dilaton finding its way to a precarious weak coupling minimum is made irrelevant by having eternal modular inflation at the vacuum supersymmetry breaking scale after the dilaton is stabilised. Neither this eternal inflation nor the subsequent non-slow-roll modular inflation destabilise the dilaton from its precarious minimum due to the low energy scale of the inflation and consequent small back reaction on the dilaton potential. The observed flat CMB spectrum is obtained from fluctuations in the angular component of a modulus near a symmetric point, which are hugely magnified by the roll down of the modulus to Planckian values, allowing them to dominate the final curvature perturbation. We also give precise calculations of the spectral index and its running.

  9. Biosynthesis of the Allylmalonyl-CoA Extender Unit for the FK506 Polyketide Synthase (PKS) Proceeds Through a Dedicated PKS and Facilitates the Mutasynthesis of Novel Analogs

    PubMed Central

    Mo, SangJoon; Kim, Dong Hwan; Lee, Jong Hyun; Park, Je Won; Basnet, Devi B.; Ban, Yeon Hee; Yoo, Young Ji; Chen, Shu-wei; Park, Sung Ryeol; Choi, Eun Ae; Kim, Eunji; Jin, Ying-Yu; Lee, Sung-Kwon; Park, Ju Yeol; Liu, Yuan; Lee, Mi Ok; Lee, Keum Soon; Kim, Sang Jun; Kim, Dooil; Park, Byoung Chul; Lee, Sang-gi; Kwon, Ho Jeong; Suh, Joo-Won; Moore, Bradley S.; Lim, Si-Kyu; Yoon, Yeo Joon

    2011-01-01

    The allyl moiety of the immunosuppressive agent FK506 is structurally unique amongst polyketides and critical for its potent biological activity. Here, we detail the biosynthetic pathway to allylmalonyl-coenzyme A (CoA), from which the FK506 allyl group is derived, based on a comprehensive chemical, biochemical and genetic interrogation of three FK506 gene clusters. A discrete polyketide synthase (PKS) with noncanonical domain architecture presumably in coordination with the fatty acid synthase pathway of the host catalyzes a multi-step enzymatic reaction to allylmalonyl-CoA via trans-2-pentenyl-acyl carrier protein. Characterization of this discrete pathway facilitated the engineered biosynthesis of novel allyl group-modified FK506 analogs, namely 36-fluoro-FK520 and 36-methyl-FK506, the latter of which exhibits improved neurite outgrowth activity. This unique feature of FK506 biosynthesis, in which a dedicated PKS provides an atypical extender unit for the main modular PKS, illuminates a new strategy for the combinatorial biosynthesis of designer macrolide scaffolds as well as FK506 analogs. PMID:21175203

  10. A Modular Robotic Architecture

    DTIC Science & Technology

    1990-11-01

    DATES COVERED AD-A232 007 Januar 1991 professional paper5 FUNOING NUMBERS A MODULAR ROBOTIC ARCHITECTURE PR: ZE92 WU: DN300029 PE: 0602936N - S. AUTHOR...mobile robots will help alleviate these problems, and, if made widely available, will promote standardization and compatibility among systems throughout...the industry. The Modular Robotic Architecture (MRA) is a generic control system that meets the above needs by providing developers with a standard set

  11. Modularity and mental architecture.

    PubMed

    Robbins, Philip

    2013-11-01

    Debates about the modularity of cognitive architecture have been ongoing for at least the past three decades, since the publication of Fodor's landmark book The Modularity of Mind. According to Fodor, modularity is essentially tied to informational encapsulation, and as such is only found in the relatively low-level cognitive systems responsible for perception and language. According to Fodor's critics in the evolutionary psychology camp, modularity simply reflects the fine-grained functional specialization dictated by natural selection, and it characterizes virtually all aspects of cognitive architecture, including high-level systems for judgment, decision making, and reasoning. Though both of these perspectives on modularity have garnered support, the current state of evidence and argument suggests that a broader skepticism about modularity may be warranted. WIREs Cogn Sci 2013, 4:641-649. doi: 10.1002/wcs.1255 CONFLICT OF INTEREST: The author has declared no conflicts of interest for this article. For further resources related to this article, please visit the WIREs website.

  12. Identification of olivetolic acid cyclase from Cannabis sativa reveals a unique catalytic route to plant polyketides

    PubMed Central

    Gagne, Steve J.; Stout, Jake M.; Liu, Enwu; Boubakir, Zakia; Clark, Shawn M.; Page, Jonathan E.

    2012-01-01

    Δ9-Tetrahydrocannabinol (THC) and other cannabinoids are responsible for the psychoactive and medicinal properties of Cannabis sativa L. (marijuana). The first intermediate in the cannabinoid biosynthetic pathway is proposed to be olivetolic acid (OA), an alkylresorcinolic acid that forms the polyketide nucleus of the cannabinoids. OA has been postulated to be synthesized by a type III polyketide synthase (PKS) enzyme, but so far type III PKSs from cannabis have been shown to produce catalytic byproducts instead of OA. We analyzed the transcriptome of glandular trichomes from female cannabis flowers, which are the primary site of cannabinoid biosynthesis, and searched for polyketide cyclase-like enzymes that could assist in OA cyclization. Here, we show that a type III PKS (tetraketide synthase) from cannabis trichomes requires the presence of a polyketide cyclase enzyme, olivetolic acid cyclase (OAC), which catalyzes a C2–C7 intramolecular aldol condensation with carboxylate retention to form OA. OAC is a dimeric α+β barrel (DABB) protein that is structurally similar to polyketide cyclases from Streptomyces species. OAC transcript is present at high levels in glandular trichomes, an expression profile that parallels other cannabinoid pathway enzymes. Our identification of OAC both clarifies the cannabinoid pathway and demonstrates unexpected evolutionary parallels between polyketide biosynthesis in plants and bacteria. In addition, the widespread occurrence of DABB proteins in plants suggests that polyketide cyclases may play an overlooked role in generating plant chemical diversity. PMID:22802619

  13. Identification of olivetolic acid cyclase from Cannabis sativa reveals a unique catalytic route to plant polyketides.

    PubMed

    Gagne, Steve J; Stout, Jake M; Liu, Enwu; Boubakir, Zakia; Clark, Shawn M; Page, Jonathan E

    2012-07-31

    Δ(9)-Tetrahydrocannabinol (THC) and other cannabinoids are responsible for the psychoactive and medicinal properties of Cannabis sativa L. (marijuana). The first intermediate in the cannabinoid biosynthetic pathway is proposed to be olivetolic acid (OA), an alkylresorcinolic acid that forms the polyketide nucleus of the cannabinoids. OA has been postulated to be synthesized by a type III polyketide synthase (PKS) enzyme, but so far type III PKSs from cannabis have been shown to produce catalytic byproducts instead of OA. We analyzed the transcriptome of glandular trichomes from female cannabis flowers, which are the primary site of cannabinoid biosynthesis, and searched for polyketide cyclase-like enzymes that could assist in OA cyclization. Here, we show that a type III PKS (tetraketide synthase) from cannabis trichomes requires the presence of a polyketide cyclase enzyme, olivetolic acid cyclase (OAC), which catalyzes a C2-C7 intramolecular aldol condensation with carboxylate retention to form OA. OAC is a dimeric α+β barrel (DABB) protein that is structurally similar to polyketide cyclases from Streptomyces species. OAC transcript is present at high levels in glandular trichomes, an expression profile that parallels other cannabinoid pathway enzymes. Our identification of OAC both clarifies the cannabinoid pathway and demonstrates unexpected evolutionary parallels between polyketide biosynthesis in plants and bacteria. In addition, the widespread occurrence of DABB proteins in plants suggests that polyketide cyclases may play an overlooked role in generating plant chemical diversity.

  14. Molecular Evolution of Aromatic Polyketides and Comparative Sequence Analysis of Polyketide Ketosynthase and 16S Ribosomal DNA Genes from Various Streptomyces Species

    PubMed Central

    Metsä-Ketelä, Mikko; Halo, Laura; Munukka, Eveliina; Hakala, Juha; Mäntsälä, Pekka; Ylihonko, Kristiina

    2002-01-01

    A 613-bp fragment of an essential ketosynthase gene from the biosynthetic pathway of aromatic polyketide antibiotics was sequenced from 99 actinomycetes isolated from soil. Phylogenetic analysis showed that the isolates clustered into clades that correspond to the various classes of aromatic polyketides. Additionally, sequencing of a 120-bp fragment from the γ-variable region of 16S ribosomal DNA (rDNA) and subsequent comparative sequence analysis revealed incongruity between the ketosynthase and 16S rDNA phylogenetic trees, which strongly suggests that there has been horizontal transfer of aromatic polyketide biosynthesis genes. The results show that the ketosynthase tree could be used for DNA fingerprinting of secondary metabolites and for screening interesting aromatic polyketide biosynthesis genes. Furthermore, the movement of the ketosynthase genes suggests that traditional marker molecules like 16S rDNA give misleading information about the biosynthesis potential of aromatic polyketides, and thus only molecules that are directly involved in the biosynthesis of secondary metabolites can be used to gain information about the biodiversity of antibiotic production in different actinomycetes. PMID:12200302

  15. Recent advances in genome-based polyketide discovery.

    PubMed

    Helfrich, Eric J N; Reiter, Silke; Piel, Jörn

    2014-10-01

    Polyketides are extraordinarily diverse secondary metabolites of great pharmacological value and with interesting ecological functions. The post-genomics era has led to fundamental changes in natural product research by inverting the workflow of secondary metabolite discovery. As opposed to traditional bioactivity-guided screenings, genome mining is an in silico method to screen and analyze sequenced genomes for natural product biosynthetic gene clusters. Since genes for known compounds can be recognized at the early computational stage, genome mining presents an opportunity for dereplication. This review highlights recent progress in bioinformatics, pathway engineering and chemical analytics to extract the biosynthetic secrets hidden in the genome of both well-known natural product sources as well as previously neglected bacteria.

  16. Prenylated phenyl polyketides and acylphloroglucinols from Hypericum peplidifolium.

    PubMed

    Fobofou, Serge Alain Tanemossu; Harmon, Chelsea Rebecca; Lonfouo, Antoine Honoré Nkuete; Franke, Katrin; Wright, Stephen M; Wessjohann, Ludger A

    2016-04-01

    In search for new or chemo-taxonomically relevant bioactive compounds from chemically unexplored Hypericum species, four previously undescribed natural products, named peplidiforones A-D were isolated and characterized from Hypericum peplidifolium A. Rich., together with six known compounds. The structures of all compounds were elucidated by extensive 1D- and 2D-NMR experiments, high resolution mass spectrometric analyses (HR-MS), and by comparison with data reported in the literature. Seven of these compounds are phenyl polyketides while three are acylphloroglucinol type compounds. Peplidiforone C, which possesses an unusual carbon skeleton consisting of a furan ring substituted by a 2,2-dimethylbut-3-enoyl moiety, is the first example of a prenylated furan derivative isolated from the genus Hypericum. The cytotoxicity, antifungal, and anti-herpes simplex virus type 1 (HSV-1) activities of extracts and compounds are described.

  17. Mycolactones: immunosuppressive and cytotoxic polyketides produced by aquatic mycobacteria

    PubMed Central

    Hong, Hui; Demangel, Caroline; Pidot, Sacha J.; Leadlay, Peter F.

    2008-01-01

    Mycolactones are a family of highly related macrocyclic polyketides that exhibit immunosuppressive and cytotoxic properties. First discovered in 1999, they are the primary virulence factors produced by the environmental human pathogen Mycobacterium ulcerans, the causative agent of Buruli ulcer, and by some closely-related aquatic mycobacteria that cause disease in fish and frogs. Mycolactones are characterized by a common 12-membered lactone core to which is appended an unsaturated fatty acyl side-chain of variable length and oxidation state. This Highlight summarizes recent progress in understanding the structural diversity of the mycolactones, their biological activity and mode of action in mammalian cells, and the genetics, evolution, and enzymology of their biosynthesis. PMID:18497894

  18. Exploiting the genetic potential of polyketide producing streptomycetes.

    PubMed

    Weber, T; Welzel, K; Pelzer, S; Vente, A; Wohlleben, W

    2003-12-19

    Streptomycetes are the most important bacterial producers of bioactive secondary metabolites such as antibiotics or cytostatics. Due to the emerging resistance of pathogenic bacteria to all commonly used antibiotics, new and modified natural compounds are required for the development of novel drugs. In addition to the classical screening for natural compounds, genome driven approaches like combinatorial biosynthesis are permanently gaining relevance for the generation of new structures. This technology utilizes the combination of genes from different biosynthesis pathways resulting in the production of novel or modified metabolites. The basis for this strategy is the access to a significant number of genes and the knowledge about the activity and specificity of the enzymes encoded by them. A joint initiative was started to exploit the biosynthesis gene clusters from streptomycetes. In this publication, an overview of the strategy for the identification and characterization of numerous biosynthesis gene clusters for polyketides displaying interesting functions and particular structural features is given.

  19. Identification of Unique Type II Polyketide Synthase Genes in Soil

    PubMed Central

    Wawrik, Boris; Kerkhof, Lee; Zylstra, Gerben J.; Kukor, Jerome J.

    2005-01-01

    Many bacteria, particularly actinomycetes, are known to produce secondary metabolites synthesized by polyketide synthases (PKS). Bacterial polyketides are a particularly rich source of bioactive molecules, many of which are of potential pharmaceutical relevance. To directly access PKS gene diversity from soil, we developed degenerate PCR primers for actinomycete type II KSα (ketosynthase) genes. Twenty-one soil samples were collected from diverse sources in New Jersey, and their bacterial communities were compared by terminal restriction fragment length polymorphism (TRFLP) analysis of PCR products generated using bacterial 16S rRNA gene primers (27F and 1525R) as well as an actinomycete-specific forward primer. The distribution of actinomycetes was highly variable but correlated with the overall bacterial species composition as determined by TRFLP. Two samples were identified to contain a particularly rich and unique actinomycete community based on their TRFLP patterns. The same samples also contained the greatest diversity of KSα genes as determined by TRFLP analysis of KSα PCR products. KSα PCR products from these and three additional samples with interesting TRFLP pattern were cloned, and seven novel clades of KSα genes were identified. Greatest sequence diversity was observed in a sample containing a moderate number of peaks in its KSα TRFLP. The nucleotide sequences were between 74 and 81% identical to known sequences in GenBank. One cluster of sequences was most similar to the KSα involved in ardacin (glycopeptide antibiotic) production by Kibdelosporangium aridum. The remaining sequences showed greatest similarity to the KSα genes in pathways producing the angucycline-derived antibiotics simocyclinone, pradimicin, and jasomycin. PMID:15870305

  20. Construction of a Part of a 3-Hydroxypropionate Cycle for Heterologous Polyketide Biosynthesis in Escherichia coli

    SciTech Connect

    Yuzawa, S; Chiba, N; Katz, L; Keasling, JD

    2012-12-11

    Polyketides, an important class of natural products with, complex chemical structures, are widely used as antibiotics and other pharmaceutical agents. A clear barrier to heterologous polyketide biosynthesis in Escherichia coli is the lack of (2S)-methylmalonyl-CoA, a common substrate of multimodular polyketide synthases. Here we report a route for synthesizing (2S)-methylmalonyl-CoA from malonyl-CoA with a 3-hydroxypropionate cycle in thermoacidophilic crenarchaeon. The engineered E. coli strain produced both propionyl-CoA and methylmalonyl-CoA at intracellular levels similar to those of acetyl-CoA and succinyl-CoA, respectively. This approach may open a way to produce a variety of polyketide drugs in E. coli from renewable carbon sources.

  1. Use of a biosynthetic intermediate to explore the chemical diversity of pseudo-natural fungal polyketides

    NASA Astrophysics Data System (ADS)

    Asai, Teigo; Tsukada, Kento; Ise, Satomi; Shirata, Naoki; Hashimoto, Makoto; Fujii, Isao; Gomi, Katsuya; Nakagawara, Kosuke; Kodama, Eiichi N.; Oshima, Yoshiteru

    2015-09-01

    The structural complexity and diversity of natural products make them attractive sources for potential drug discovery, with their characteristics being derived from the multi-step combination of enzymatic and non-enzymatic conversions of intermediates in each biosynthetic pathway. Intermediates that exhibit multipotent behaviour have great potential for use as starting points in diversity-oriented synthesis. Inspired by the biosynthetic pathways that form complex metabolites from simple intermediates, we developed a semi-synthetic process that combines heterologous biosynthesis and artificial diversification. The heterologous biosynthesis of fungal polyketide intermediates led to the isolation of novel oligomers and provided evidence for ortho-quinonemethide equivalency in their isochromene form. The intrinsic reactivity of the isochromene polyketide enabled us to access various new chemical entities by modifying and remodelling the polyketide core and through coupling with indole molecules. We thus succeeded in generating exceptionally diverse pseudo-natural polyketides through this process and demonstrated an advanced method of using biosynthetic intermediates.

  2. Fumonisin-nonproducing mutants exhibit differential expression of putative polyketide biosynthetic gene clusters in Fusarium verticillioides

    USDA-ARS?s Scientific Manuscript database

    The maize pathogen Fusarium verticillioides produces a group of polyketide derived secondary metabolites called fumonisins. Fumonisins can cause diseases in animals, and have been correlated epidemiologically with esophageal cancer and birth defects in humans. The fumonisin biosynthetic gene clust...

  3. Symmetric modular torsatron

    DOEpatents

    Rome, J.A.; Harris, J.H.

    1984-01-01

    A fusion reactor device is provided in which the magnetic fields for plasma confinement in a toroidal configuration is produced by a plurality of symmetrical modular coils arranged to form a symmetric modular torsatron referred to as a symmotron. Each of the identical modular coils is helically deformed and comprise one field period of the torsatron. Helical segments of each coil are connected by means of toroidally directed windbacks which may also provide part of the vertical field required for positioning the plasma. The stray fields of the windback segments may be compensated by toroidal coils. A variety of magnetic confinement flux surface configurations may be produced by proper modulation of the winding pitch of the helical segments of the coils, as in a conventional torsatron, winding the helix on a noncircular cross section and varying the poloidal and radial location of the windbacks and the compensating toroidal ring coils.

  4. Self Evolving Modular Network

    NASA Astrophysics Data System (ADS)

    Tokunaga, Kazuhiro; Kawabata, Nobuyuki; Furukawa, Tetsuo

    We propose a novel modular network called the Self-Evolving Modular Network (SEEM). The SEEM has a modular network architecture with a graph structure and these following advantages: (1) new modules are added incrementally to allow the network to adapt in a self-organizing manner, and (2) graph's paths are formed based on the relationships between the models represented by modules. The SEEM is expected to be applicable to evolving functions of an autonomous robot in a self-organizing manner through interaction with the robot's environment and categorizing large-scale information. This paper presents the architecture and an algorithm for the SEEM. Moreover, performance characteristic and effectiveness of the network are shown by simulations using cubic functions and a set of 3D-objects.

  5. Modular optical detector system

    DOEpatents

    Horn, Brent A.; Renzi, Ronald F.

    2006-02-14

    A modular optical detector system. The detector system is designed to detect the presence of molecules or molecular species by inducing fluorescence with exciting radiation and detecting the emitted fluorescence. Because the system is capable of accurately detecting and measuring picomolar concentrations it is ideally suited for use with microchemical analysis systems generally and capillary chromatographic systems in particular. By employing a modular design, the detector system provides both the ability to replace various elements of the detector system without requiring extensive realignment or recalibration of the components as well as minimal user interaction with the system. In addition, the modular concept provides for the use and addition of a wide variety of components, including optical elements (lenses and filters), light sources, and detection means, to fit particular needs.

  6. Cystochromones, Unusual Chromone-Containing Polyketides from the Myxobacterium Cystobacter sp. MCy9104.

    PubMed

    Nadmid, Suvd; Plaza, Alberto; Garcia, Ronald; Müller, Rolf

    2015-08-28

    Seven new chromone-containing polyketides, termed cystochromones A-G, were isolated from the myxobacterial strain Cystobacter sp. MCy9104. Their structures were elucidated using comprehensive NMR spectroscopy and HR-MS/MS. Cystochromones bear a pentadecyl moiety unusually attached at C-5 of the chromone ring. Moreover, isotope-labeled substrate feeding experiments and NMR analysis suggested a hybrid iso-fatty acid and polyketide synthase biosynthetic pathway for these secondary metabolites.

  7. The toxic dinoflagellate Karenia brevis encodes novel type I-like polyketide synthases containing discrete catalytic domains.

    PubMed

    Monroe, Emily A; Van Dolah, Frances M

    2008-07-01

    Karenia brevis is the Florida red tide dinoflagellate responsible for detrimental effects on human and environmental health through the production of brevetoxins. Brevetoxins are thought to be synthesized by a polyketide synthase (PKS) complex, but the gene cluster for this PKS has yet to be identified. Here, eight PKS transcripts were identified in K. brevis by high throughput cDNA library screening. Full length sequences were obtained through 3' and 5' RACE, which demonstrated the presence of polyadenylation, 3'-UTRs, and an identical dinoflagellate-specific spliced leader sequence at the 5' end of PKS transcripts. Six transcripts encoded for individual ketosynthase (KS) domains, one ketoreductase (KR), and one transcript encoded both acyl carrier protein (ACP) and KS domains. Transcript lengths ranged from 1875 to 3397 nucleotides, based on sequence analysis, and were confirmed by northern blotting. Baysian phylogenetic analysis of the K. brevis KS domains placed them well within the protist type I PKS clade. Thus although most similar to type I modular PKSs, the presence of individual catalytic domains on separate transcripts suggests a protein structure more similar to type II PKSs, in which each catalytic domain resides on an individual protein. These results identify an unprecedented PKS structure in a toxic dinoflagellate.

  8. Modular biowaste monitoring system

    NASA Technical Reports Server (NTRS)

    Fogal, G. L.

    1975-01-01

    The objective of the Modular Biowaste Monitoring System Program was to generate and evaluate hardware for supporting shuttle life science experimental and diagnostic programs. An initial conceptual design effort established requirements and defined an overall modular system for the collection, measurement, sampling and storage of urine and feces biowastes. This conceptual design effort was followed by the design, fabrication and performance evaluation of a flight prototype model urine collection, volume measurement and sampling capability. No operational or performance deficiencies were uncovered as a result of the performance evaluation tests.

  9. Modular Optofluidic Systems (MOPS)

    NASA Astrophysics Data System (ADS)

    Ackermann, Tobias N.; Dietvorst, Jiri; Sanchis, Ana; Salvador, Juan P.; Munoz-Berbel, Xavier; Alvarez-Conde, Erica; Kopp, Daniel; Zappe, Hans; Marco, M.-Pilar; Llobera, Andreu

    2016-12-01

    Elementary PDMS-based building blocks of fluidic, optical and optofluidic components for Lab on a chip (LOC) platforms has here been developed. All individual modules are compatible and can be anchored and released with the help of puzzle-type connectors This approach is a powerful toolbox to create modular optofluidic systems (MOPS), which can be modified/upgraded to user needs and in-situ reconfigurable. In addition, the PDMS can locally be functionalized, defining a modular biosensor. Measurements in absorbance and fluorescence have been pursued as demonstrator.

  10. Modular total absorption spectrometer

    NASA Astrophysics Data System (ADS)

    Karny, M.; Rykaczewski, K. P.; Fijałkowska, A.; Rasco, B. C.; Wolińska-Cichocka, M.; Grzywacz, R. K.; Goetz, K. C.; Miller, D.; Zganjar, E. F.

    2016-11-01

    The design and performance of the Modular Total Absorption Spectrometer built and commissioned at the Oak Ridge National Laboratory is presented. The active volume of the detector is approximately one ton of NaI(Tl), which results in very high full γ energy peak efficiency of 71% at 6 MeV and nearly flat efficiency of around 81.5% for low energy γ-rays between 300 keV and 1 MeV. In addition to the high peak efficiency, the modular construction of the detector permits the use of a γ-coincidence technique in data analysis as well as β-delayed neutron observation.

  11. The LINKS motif zippers trans-acyltransferase polyketide synthase assembly lines into a biosynthetic megacomplex

    PubMed Central

    Gay, Darren C.; Wagner, Drew T.; Meinke, Jessica L.; Zogzas, Charles E.; Gay, Glen R.; Keatinge-Clay, Adrian T.

    2016-01-01

    Polyketides such as the clinically-valuable antibacterial agent mupirocin are constructed by architecturally-sophisticated assembly lines known as trans-acyltransferase polyketide synthases. Organelle-sized megacomplexes composed of several copies of trans-acyltransferase polyketide synthase assembly lines have been observed by others through transmission electron microscopy to be located at the Bacillus subtilis plasma membrane, where the synthesis and export of the antibacterial polyketide bacillaene takes place. In this work we analyze ten crystal structures of trans-acyltransferase polyketide synthases ketosynthase domains, seven of which are reported here for the first time, to characterize a motif capable of zippering assembly lines into a megacomplex. While each of the three-helix LINKS (Laterally-INteracting Ketosynthase Sequence) motifs is observed to similarly dock with a spatially-reversed copy of itself through hydrophobic and ionic interactions, the amino acid sequences of this motif are not conserved. Such a code is appropriate for mediating homotypic contacts between assembly lines to ensure the ordered self-assembly of a noncovalent, yet tightly-knit, enzymatic network. LINKS-mediated lateral interactions would also have the effect of bolstering the vertical association of the polypeptides that comprise a polyketide synthase assembly line. PMID:26724270

  12. Inhibition Kinetics And Emodin Cocrystal Structure of a Type II Polyketide Ketoreductase

    SciTech Connect

    Korman, T.P.; Tan, Y.-H.; Wong, J.; Luo, R.; Tsai, S.-C.

    2009-05-20

    Type II polyketides are a class of natural products that include pharmaceutically important aromatic compounds such as the antibiotic tetracycline and antitumor compound doxorubicin. The type II polyketide synthase (PKS) is a complex consisting of 5-10 standalone domains homologous to fatty acid synthase (FAS). Polyketide ketoreductase (KR) provides regio- and stereochemical diversity during the reduction. How the type II polyketide KR specifically reduces only the C9 carbonyl group is not well understood. The cocrystal structures of actinorhodin polyketide ketoreductase (actKR) bound with NADPH or NADP{sup +} and the inhibitor emodin were solved with the wild type and P94L mutant of actKR, revealing the first observation of a bent p-quinone in an enzyme active site. Molecular dynamics simulation help explain the origin of the bent geometry. Extensive screening for in vitro substrates shows that unlike FAS KR, the actKR prefers bicyclic substrates. Inhibition kinetics indicate that actKR follows an ordered Bi Bi mechanism. Together with docking simulations that identified a potential phosphopantetheine binding groove, the structural and functional studies reveal that the C9 specificity is a result of active site geometry and substrate ring constraints. The results lay the foundation for the design of novel aromatic polyketide natural products with different reduction patterns.

  13. Saccharomyces cerevisiae as a tool for mining, studying and engineering fungal polyketide synthases.

    PubMed

    Bond, Carly; Tang, Yi; Li, Li

    2016-04-01

    Small molecule secondary metabolites produced by organisms such as plants, bacteria, and fungi form a fascinating and important group of natural products, many of which have shown promise as medicines. Fungi in particular have been important sources of natural product polyketide pharmaceuticals. While the structural complexity of these polyketides makes them interesting and useful bioactive compounds, these same features also make them difficult and expensive to prepare and scale-up using synthetic methods. Currently, nearly all commercial polyketides are prepared through fermentation or semi-synthesis. However, elucidation and engineering of polyketide pathways in the native filamentous fungi hosts are often hampered due to a lack of established genetic tools and of understanding of the regulation of fungal secondary metabolisms. Saccharomyces cerevisiae has many advantages beneficial to the study and development of polyketide pathways from filamentous fungi due to its extensive genetic toolbox and well-studied metabolism. This review highlights the benefits S. cerevisiae provides as a tool for mining, studying, and engineering fungal polyketide synthases (PKSs), as well as notable insights this versatile tool has given us into the mechanisms and products of fungal PKSs.

  14. Saccharomyces cerevisiae as a tool for mining, studying and engineering fungal polyketide synthases

    PubMed Central

    Bond, Carly; Tang, Yi; Li, Li

    2016-01-01

    Small molecule secondary metabolites produced by organisms such as plants, bacteria, and fungi form a fascinating and important group of natural products, many of which have shown promise as medicines. Fungi in particular have been important sources of natural product polyketide pharmaceuticals. While the structural complexity of these polyketides makes them interesting and useful bioactive compounds, these same features also make them difficult and expensive to prepare and scale-up using synthetic methods. Currently, nearly all commercial polyketides are prepared through fermentation or semi-synthesis. However, elucidation and engineering of polyketide pathways in the native filamentous fungi hosts are often hampered due to a lack of established genetic tools and of understanding of the regulation of fungal secondary metabolisms. Saccharomyces cerevisiae has many advantages beneficial to the study and development of polyketide pathways from filamentous fungi due to its extensive genetic toolbox and well-studied metabolism. This review highlights the benefits S. cerevisiae provides as a tool for mining, studying, and engineering fungal polyketide synthases (PKSs), as well as notable insights this versatile tool has given us into the mechanisms and products of fungal PKSs. PMID:26850128

  15. Modular invariant inflation

    SciTech Connect

    Kobayashi, Tatsuo; Nitta, Daisuke; Urakawa, Yuko

    2016-08-08

    Modular invariance is a striking symmetry in string theory, which may keep stringy corrections under control. In this paper, we investigate a phenomenological consequence of the modular invariance, assuming that this symmetry is preserved as well as in a four dimensional (4D) low energy effective field theory. As a concrete setup, we consider a modulus field T whose contribution in the 4D effective field theory remains invariant under the modular transformation and study inflation drived by T. The modular invariance restricts a possible form of the scalar potenntial. As a result, large field models of inflation are hardly realized. Meanwhile, a small field model of inflation can be still accomodated in this restricted setup. The scalar potential traced during the slow-roll inflation mimics the hilltop potential V{sub ht}, but it also has a non-negligible deviation from V{sub ht}. Detecting the primordial gravitational waves predicted in this model is rather challenging. Yet, we argue that it may be still possible to falsify this model by combining the information in the reheating process which can be determined self-completely in this setup.

  16. Modular invariant gaugino condensation

    SciTech Connect

    Gaillard, M.K.

    1991-05-09

    The construction of effective supergravity lagrangians for gaugino condensation is reviewed and recent results are presented that are consistent with modular invariance and yield a positive definite potential of the noscale type. Possible implications for phenomenology are briefly discussed. 29 refs.

  17. Modular invariant inflation

    NASA Astrophysics Data System (ADS)

    Kobayashi, Tatsuo; Nitta, Daisuke; Urakawa, Yuko

    2016-08-01

    Modular invariance is a striking symmetry in string theory, which may keep stringy corrections under control. In this paper, we investigate a phenomenological consequence of the modular invariance, assuming that this symmetry is preserved as well as in a four dimensional (4D) low energy effective field theory. As a concrete setup, we consider a modulus field T whose contribution in the 4D effective field theory remains invariant under the modular transformation and study inflation drived by T. The modular invariance restricts a possible form of the scalar potenntial. As a result, large field models of inflation are hardly realized. Meanwhile, a small field model of inflation can be still accomodated in this restricted setup. The scalar potential traced during the slow-roll inflation mimics the hilltop potential Vht, but it also has a non-negligible deviation from Vht. Detecting the primordial gravitational waves predicted in this model is rather challenging. Yet, we argue that it may be still possible to falsify this model by combining the information in the reheating process which can be determined self-completely in this setup.

  18. A Modular CAI System.

    ERIC Educational Resources Information Center

    Van Der Mast, Charles

    The experimental CAI system which is being tested at Delft University of Technology is structured in a modular manner to account for high changeability. The concept formulated for this project was the outcome of research into technological, organizational, and educational developments in CAI, and the enumeration of the common aspects of the…

  19. Modular core holder

    SciTech Connect

    Mueller, J.; Cole, C.W.; Hamid, S.; Lucas, J.K.

    1991-03-05

    This patent describes a modular core holder. It comprises: a sleeve, forming an internal cavity for receiving a core. The sleeve including segments; support means, overlying the sleeve, for supporting the sleeve; and access means, positioned between at least two of the segments of the sleeve, for allowing measurement of conditions within the internal cavity.

  20. MRV - Modular Robotic Vehicle

    NASA Technical Reports Server (NTRS)

    Ridley, Justin; Bluethmann, Bill

    2015-01-01

    The Modular Robotic Vehicle, or MRV, completed in 2013, was developed at the Johnson Space Center in order to advance technologies which have applications for future vehicles both in space and on Earth. With seating for two people, MRV is a fully electric vehicle modeled as a "city car", suited for busy urban environments.

  1. Modular Perspectives on Bilingualism.

    ERIC Educational Resources Information Center

    Francis, Norbert

    2002-01-01

    This research review traces the current discussion on models of bilingualism to the contributions of Vygotsky and Luria. Proposes that a modular approach to studying the different aspects of bilingual development promises to chart a course toward finding a broader common ground around research findings and interpretations that appear to be…

  2. Modularity in robotic systems

    NASA Technical Reports Server (NTRS)

    Tesar, Delbert; Butler, Michael S.

    1989-01-01

    Most robotic systems today are designed one at a time, at a high cost of time and money. This wasteful approach has been necessary because the industry has not established a foundation for the continued evolution of intelligent machines. The next generation of robots will have to be generic, versatile machines capable of absorbing new technology rapidly and economically. This approach is demonstrated in the success of the personal computer, which can be upgraded or expanded with new software and hardware at virtually every level. Modularity is perceived as a major opportunity to reduce the 6 to 7 year design cycle time now required for new robotic manipulators, greatly increasing the breadth and speed of diffusion of robotic systems in manufacturing. Modularity and its crucial role in the next generation of intelligent machines are the focus of interest. The main advantages that modularity provides are examined; types of modules needed to create a generic robot are discussed. Structural modules designed by the robotics group at the University of Texas at Austin are examined to demonstrate the advantages of modular design.

  3. Modular, Multilayer Perceptron

    NASA Technical Reports Server (NTRS)

    Cheng, Li-Jen; Liu, Tsuen-Hsi

    1991-01-01

    Combination of proposed modular, multilayer perceptron and algorithm for its operation recognizes new objects after relatively brief retraining sessions. (Perceptron is multilayer, feedforward artificial neural network fully connected and trained via back-propagation learning algorithm.) Knowledge pertaining to each object to be recognized resides in subnetwork of full network, therefore not necessary to retrain full network to recognize each new object.

  4. Identification and Biotechnological Application of Novel Regulatory Genes Involved in Streptomyces Polyketide Overproduction through Reverse Engineering Strategy

    PubMed Central

    Nah, Ji-Hye; Kim, Hye-Jin; Lee, Han-Na; Lee, Mi-Jin; Choi, Si-Sun; Kim, Eung-Soo

    2013-01-01

    Polyketide belongs to a family of abundant natural products typically produced by the filamentous soil bacteria Streptomyces. Similar to the biosynthesis of most secondary metabolites produced in the Streptomyces species, polyketide compounds are synthesized through tight regulatory networks in the cell, and thus extremely low levels of polyketides are typically observed in wild-type strains. Although many Streptomyces polyketides and their derivatives have potential to be used as clinically important pharmaceutical drugs, traditional strain improvement strategies such as random recursive mutagenesis have long been practiced with little understanding of the molecular basis underlying enhanced polyketide production. Recently, identifying, understanding, and applying a novel polyketide regulatory system identified from various Omics approaches, has become an important tool for rational Streptomyces strain improvement. In this paper, DNA microarray-driven reverse engineering efforts for improving titers of polyketides are briefly summarized, primarily focusing on our recent results of identification and application of novel global regulatory genes such as wblA, SCO1712, and SCO5426 in Streptomyces species. Sequential targeted gene manipulation involved in polyketide biosynthetic reguation synergistically provided an efficient and rational strategy for Streptomyces strain improvement. Moreover, the engineered regulation-optimized Streptomyces mutant strain was further used as a surrogate host for heterologous expression of polyketide pathway. PMID:23555090

  5. Metabolic engineering of Streptomyces venezuelae for malonyl-CoA biosynthesis to enhance heterologous production of polyketides.

    PubMed

    Maharjan, Sushila; Park, Je Won; Yoon, Yeo Joon; Lee, Hei Chan; Sohng, Jae Kyung

    2010-02-01

    Using metabolic engineering, we developed Streptomyces venezuelae YJ028 as an efficient heterologous host to increase the malonyl-CoA pool to be directed towards enhanced production of various polyketides. To probe the applicability of newly developed hosts in the heterologous production of polyketides, we expressed type III polyketide synthase, 1,3,6,8-tetrahydroxynaphthalene synthase, in these hosts. Flaviolin production was doubled by expression of acetyl-CoA carboxylase (ACCase) and 4-fold by combined expression of ACCase, metK1-sp and afsR-sp. Thus, the newly developed Streptomyces venezuelae YJ028 hosts produce heterologous polyketides more efficiently than the parent strain.

  6. C-S bond cleavage by a polyketide synthase domain

    PubMed Central

    Ma, Ming; Lohman, Jeremy R.; Liu, Tao; Shen, Ben

    2015-01-01

    Leinamycin (LNM) is a sulfur-containing antitumor antibiotic featuring an unusual 1,3-dioxo-1,2-dithiolane moiety that is spiro-fused to a thiazole-containing 18-membered lactam ring. The 1,3-dioxo-1,2-dithiolane moiety is essential for LNM’s antitumor activity, by virtue of its ability to generate an episulfonium ion intermediate capable of alkylating DNA. We have previously cloned and sequenced the lnm gene cluster from Streptomyces atroolivaceus S-140. In vivo and in vitro characterizations of the LNM biosynthetic machinery have since established that: (i) the 18-membered macrolactam backbone is synthesized by LnmP, LnmQ, LnmJ, LnmI, and LnmG, (ii) the alkyl branch at C-3 of LNM is installed by LnmK, LnmL, LnmM, and LnmF, and (iii) leinamycin E1 (LNM E1), bearing a thiol moiety at C-3, is the nascent product of the LNM hybrid nonribosomal peptide synthetase (NRPS)-acyltransferase (AT)-less type I polyketide synthase (PKS). Sulfur incorporation at C-3 of LNM E1, however, has not been addressed. Here we report that: (i) the bioinformatics analysis reveals a pyridoxal phosphate (PLP)-dependent domain, we termed cysteine lyase (SH) domain (LnmJ-SH), within PKS module-8 of LnmJ; (ii) the LnmJ-SH domain catalyzes C-S bond cleavage by using l-cysteine and l-cysteine S-modified analogs as substrates through a PLP-dependent β-elimination reaction, establishing l-cysteine as the origin of sulfur at C-3 of LNM; and (iii) the LnmJ-SH domain, sharing no sequence homology with any other enzymes catalyzing C-S bond cleavage, represents a new family of PKS domains that expands the chemistry and enzymology of PKSs and might be exploited to incorporate sulfur into polyketide natural products by PKS engineering. PMID:26240335

  7. Biochemical characterization of the minimal polyketide synthase domains in the lovastatin nonaketide synthase LovB.

    PubMed

    Ma, Suzanne M; Tang, Yi

    2007-06-01

    The biosynthesis of lovastatin in Aspergillus terreus requires two megasynthases. The lovastatin nonaketide synthase, LovB, synthesizes the intermediate dihydromonacolin L using nine malonyl-coenzyme A molecules, and is a reducing, iterative type I polyketide synthase. The iterative type I polyketide synthase is mechanistically different from bacterial type I polyketide synthases and animal fatty acid synthases. We have cloned the minimal polyketide synthase domains of LovB as standalone proteins and assayed their activities and substrate specificities. The didomain proteins ketosynthase-malonyl-coenzyme A:acyl carrier protein acyltransferase (KS-MAT) and acyl carrier protein-condensation (ACP-CON) domain were expressed solubly in Escherichia coli. The monodomains MAT, ACP and CON were also obtained as soluble proteins. The MAT domain can be readily labeled by [1,2-(14)C]malonyl-coenzyme A and can transfer the acyl group to both the cognate LovB ACP and heterologous ACPs from bacterial type I and type II polyketide synthases. Using the LovB ACP-CON didomain as an acyl acceptor, LovB MAT transferred malonyl and acetyl groups with k(cat)/K(m) values of 0.62 min(-1).mum(-1) and 0.032 min(-1).mum(-1), respectively. The LovB MAT domain was able to substitute the Streptomyces coelicolor FabD in supporting product turnover in a bacterial type II minimal polyketide synthase assay. The activity of the KS domain was assayed independently using a KS-MAT (S656A) mutant in which the MAT domain was inactivated. The KS domain displayed no activity towards acetyl groups, but was able to recognize malonyl groups in the absence of cerulenin. The relevance of these finding to the priming mechanism of fungal polyketide synthase is discussed.

  8. 8 CFR 245a.32 - Ineligible aliens.

    Code of Federal Regulations, 2014 CFR

    2014-01-01

    ... 8 Aliens and Nationality 1 2014-01-01 2014-01-01 false Ineligible aliens. 245a.32 Section 245a.32 Aliens and Nationality DEPARTMENT OF HOMELAND SECURITY IMMIGRATION REGULATIONS ADJUSTMENT OF STATUS TO... IMMIGRATION AND NATIONALITY ACT LIFE Act Amendments Family Unity Provisions § 245a.32 Ineligible aliens. The...

  9. 8 CFR 245a.32 - Ineligible aliens.

    Code of Federal Regulations, 2012 CFR

    2012-01-01

    ... 8 Aliens and Nationality 1 2012-01-01 2012-01-01 false Ineligible aliens. 245a.32 Section 245a.32 Aliens and Nationality DEPARTMENT OF HOMELAND SECURITY IMMIGRATION REGULATIONS ADJUSTMENT OF STATUS TO... IMMIGRATION AND NATIONALITY ACT LIFE Act Amendments Family Unity Provisions § 245a.32 Ineligible aliens. The...

  10. 8 CFR 245a.32 - Ineligible aliens.

    Code of Federal Regulations, 2013 CFR

    2013-01-01

    ... 8 Aliens and Nationality 1 2013-01-01 2013-01-01 false Ineligible aliens. 245a.32 Section 245a.32 Aliens and Nationality DEPARTMENT OF HOMELAND SECURITY IMMIGRATION REGULATIONS ADJUSTMENT OF STATUS TO... IMMIGRATION AND NATIONALITY ACT LIFE Act Amendments Family Unity Provisions § 245a.32 Ineligible aliens. The...

  11. 8 CFR 245a.32 - Ineligible aliens.

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ... 8 Aliens and Nationality 1 2010-01-01 2010-01-01 false Ineligible aliens. 245a.32 Section 245a.32 Aliens and Nationality DEPARTMENT OF HOMELAND SECURITY IMMIGRATION REGULATIONS ADJUSTMENT OF STATUS TO... IMMIGRATION AND NATIONALITY ACT LIFE Act Amendments Family Unity Provisions § 245a.32 Ineligible aliens....

  12. 8 CFR 245a.32 - Ineligible aliens.

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... 8 Aliens and Nationality 1 2011-01-01 2011-01-01 false Ineligible aliens. 245a.32 Section 245a.32 Aliens and Nationality DEPARTMENT OF HOMELAND SECURITY IMMIGRATION REGULATIONS ADJUSTMENT OF STATUS TO... IMMIGRATION AND NATIONALITY ACT LIFE Act Amendments Family Unity Provisions § 245a.32 Ineligible aliens....

  13. 7 CFR 15a.32 - Housing.

    Code of Federal Regulations, 2012 CFR

    2012-01-01

    ... 7 Agriculture 1 2012-01-01 2012-01-01 false Housing. 15a.32 Section 15a.32 Agriculture Office of... § 15a.32 Housing. (a) General. A recipient shall not, on the basis of sex, apply different rules or regulations, impose different fees or requirements, or offer different services or benefits related to...

  14. 7 CFR 15a.32 - Housing.

    Code of Federal Regulations, 2014 CFR

    2014-01-01

    ... 7 Agriculture 1 2014-01-01 2014-01-01 false Housing. 15a.32 Section 15a.32 Agriculture Office of... § 15a.32 Housing. (a) General. A recipient shall not, on the basis of sex, apply different rules or regulations, impose different fees or requirements, or offer different services or benefits related to...

  15. 7 CFR 15a.32 - Housing.

    Code of Federal Regulations, 2011 CFR

    2011-01-01

    ... 7 Agriculture 1 2011-01-01 2011-01-01 false Housing. 15a.32 Section 15a.32 Agriculture Office of... § 15a.32 Housing. (a) General. A recipient shall not, on the basis of sex, apply different rules or regulations, impose different fees or requirements, or offer different services or benefits related to...

  16. 7 CFR 15a.32 - Housing.

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ... 7 Agriculture 1 2010-01-01 2010-01-01 false Housing. 15a.32 Section 15a.32 Agriculture Office of... § 15a.32 Housing. (a) General. A recipient shall not, on the basis of sex, apply different rules or regulations, impose different fees or requirements, or offer different services or benefits related to...

  17. 7 CFR 15a.32 - Housing.

    Code of Federal Regulations, 2013 CFR

    2013-01-01

    ... 7 Agriculture 1 2013-01-01 2013-01-01 false Housing. 15a.32 Section 15a.32 Agriculture Office of... § 15a.32 Housing. (a) General. A recipient shall not, on the basis of sex, apply different rules or regulations, impose different fees or requirements, or offer different services or benefits related to...

  18. Modular reflector concept study

    NASA Astrophysics Data System (ADS)

    Vaughan, D. H.

    1981-02-01

    The feasibility was studied of constructing large space structures, specifically a 100 meter paraboloidal R.F. reflector, by individually deploying a number of relatively small structural modules, and then joining them to form a single large structure in orbit. The advantage of this approach is that feasibility of a large antenna may be demonstrated by ground and flight tests of several smaller and less costly subelements. Thus, initial development costs are substantially reduced and a high degree of reliability can be obtained without commitment to construction of a very large system. The three candidate structural concepts investigated are: (1) the deployable cell module; (2) the paraboloidal extendable truss antenna adapted to modular assembly; and (3) the modular extendable truss antenna (META).

  19. Modular space station

    NASA Technical Reports Server (NTRS)

    1972-01-01

    The modular space station comprising small, shuttle-launched modules, and characterized by low initial cost and incremental manning, is described. The initial space station is designed to be delivered into orbit by three space shuttles and assembled in space. The three sections are the power/subsystems module, the crew/operations module, and the general purpose laboratory module. It provides for a crew of six. Subsequently duplicate/crew/operations and power/subsystems modules will be mated to the original modules, and provide for an additional six crewmen. A total of 17 research and applications modules is planned, three of which will be free-flying modules. Details are given on the program plan, modular characteristics, logistics, experiment support capability and requirements, operations analysis, design support analyses, and shuttle interfaces.

  20. ATIS - A modular approach

    NASA Astrophysics Data System (ADS)

    Kirson, Allan

    The author describes a modular approach to the design of an in-vehicle navigation and route guidance system that supports a phased implementation of the technology, and anticipates expected differences in implementation in different parts of the world and for different makes and models of vehicle. A series of sensors in the vehicle are used to determine the vehicle's position by dead reckoning and map-matching. The system then calculates the best route to the selected destination, taking into account the real-time traffic information received from a traffic management center, and presents route guidance instructions to the user as the route is traversed. Attention is given to modularity considerations, vehicle positioning, driver support, vehicle-to-infrastructure communications, and the role of standards.

  1. Modular reflector concept study

    NASA Technical Reports Server (NTRS)

    Vaughan, D. H.

    1981-01-01

    The feasibility was studied of constructing large space structures, specifically a 100 meter paraboloidal R.F. reflector, by individually deploying a number of relatively small structural modules, and then joining them to form a single large structure in orbit. The advantage of this approach is that feasibility of a large antenna may be demonstrated by ground and flight tests of several smaller and less costly subelements. Thus, initial development costs are substantially reduced and a high degree of reliability can be obtained without commitment to construction of a very large system. The three candidate structural concepts investigated are: (1) the deployable cell module; (2) the paraboloidal extendable truss antenna adapted to modular assembly; and (3) the modular extendable truss antenna (META).

  2. 2D NMR-spectroscopic screening reveals polyketides in ladybugs.

    PubMed

    Deyrup, Stephen T; Eckman, Laura E; McCarthy, Patrick H; Smedley, Scott R; Meinwald, Jerrold; Schroeder, Frank C

    2011-06-14

    Small molecules of biological origin continue to yield the most promising leads for drug design, but systematic approaches for exploring nature's cache of structural diversity are lacking. Here, we demonstrate the use of 2D NMR spectroscopy to screen a library of biorationally selected insect metabolite samples for partial structures indicating the presence of new chemical entities. This NMR-spectroscopic survey enabled detection of novel compounds in complex metabolite mixtures without prior fractionation or isolation. Our screen led to discovery and subsequent isolation of two families of tricyclic pyrones in Delphastus catalinae, a tiny ladybird beetle that is employed commercially as a biological pest control agent. The D. catalinae pyrones are based on 23-carbon polyketide chains forming 1,11-dioxo-2,6,10-trioxaanthracene and 4,8-dioxo-1,9,13-trioxaanthracene derivatives, representing ring systems not previously found in nature. This study highlights the utility of 2D NMR-spectroscopic screening for exploring nature's structure space and suggests that insect metabolomes remain vastly underexplored.

  3. 2D NMR-spectroscopic screening reveals polyketides in ladybugs

    PubMed Central

    Deyrup, Stephen T.; Eckman, Laura E.; McCarthy, Patrick H.; Smedley, Scott R.; Meinwald, Jerrold; Schroeder, Frank C.

    2011-01-01

    Small molecules of biological origin continue to yield the most promising leads for drug design, but systematic approaches for exploring nature’s cache of structural diversity are lacking. Here, we demonstrate the use of 2D NMR spectroscopy to screen a library of biorationally selected insect metabolite samples for partial structures indicating the presence of new chemical entities. This NMR-spectroscopic survey enabled detection of novel compounds in complex metabolite mixtures without prior fractionation or isolation. Our screen led to discovery and subsequent isolation of two families of tricyclic pyrones in Delphastus catalinae, a tiny ladybird beetle that is employed commercially as a biological pest control agent. The D. catalinae pyrones are based on 23-carbon polyketide chains forming 1,11-dioxo-2,6,10-trioxaanthracene and 4,8-dioxo-1,9,13-trioxaanthracene derivatives, representing ring systems not previously found in nature. This study highlights the utility of 2D NMR-spectroscopic screening for exploring nature’s structure space and suggests that insect metabolomes remain vastly underexplored. PMID:21646540

  4. Biosynthesis and Molecular Genetics of Polyketides in Marine Dinoflagellates

    PubMed Central

    Kellmann, Ralf; Stüken, Anke; Orr, Russell J. S.; Svendsen, Helene M.; Jakobsen, Kjetill S.

    2010-01-01

    Marine dinoflagellates are the single most important group of algae that produce toxins, which have a global impact on human activities. The toxins are chemically diverse, and include macrolides, cyclic polyethers, spirolides and purine alkaloids. Whereas there is a multitude of studies describing the pharmacology of these toxins, there is limited or no knowledge regarding the biochemistry and molecular genetics involved in their biosynthesis. Recently, however, exciting advances have been made. Expressed sequence tag sequencing studies have revealed important insights into the transcriptomes of dinoflagellates, whereas other studies have implicated polyketide synthase genes in the biosynthesis of cyclic polyether toxins, and the molecular genetic basis for the biosynthesis of paralytic shellfish toxins has been elucidated in cyanobacteria. This review summarises the recent progress that has been made regarding the unusual genomes of dinoflagellates, the biosynthesis and molecular genetics of dinoflagellate toxins. In addition, the evolution of these metabolic pathways will be discussed, and an outlook for future research and possible applications is provided. PMID:20479965

  5. Polyketide Glycosides from Bionectria ochroleuca Inhibit Candida albicans Biofilm Formation

    PubMed Central

    2015-01-01

    One of the challenges presented by Candida infections is that many of the isolates encountered in the clinic produce biofilms, which can decrease these pathogens’ susceptibilities to standard-of-care antibiotic therapies. Inhibitors of fungal biofilm formation offer a potential solution to counteracting some of the problems associated with Candida infections. A screening campaign utilizing samples from our fungal extract library revealed that a Bionectria ochroleuca isolate cultured on Cheerios breakfast cereal produced metabolites that blocked the in vitro formation of Candida albicans biofilms. A scale-up culture of the fungus was undertaken using mycobags (also known as mushroom bags or spawn bags), which afforded four known [TMC-151s C–F (1–4)] and three new [bionectriols B–D (5–7)] polyketide glycosides. All seven metabolites exhibited potent biofilm inhibition against C. albicans SC5314, as well as exerted synergistic antifungal activities in combination with amphotericin B. In this report, we describe the structure determination of the new metabolites, as well as compare the secondary metabolome profiles of fungi grown in flasks and mycobags. These studies demonstrate that mycobags offer a useful alternative to flask-based cultures for the preparative production of fungal secondary metabolites. PMID:25302529

  6. Phylogenomic and functional domain analysis of polyketide synthases in Fusarium

    SciTech Connect

    Brown, Daren W.; Butchko, Robert A.; Baker, Scott E.; Proctor, Robert H.

    2012-02-01

    Fusarium species are ubiquitous in nature, cause a range of plant diseases, and produce a variety of chemicals often referred to as secondary metabolites. Although some fungal secondary metabolites affect plant growth or protect plants from other fungi and bacteria, their presence in grain based food and feed is more often associated with a variety of diseases in plants and in animals. Many of these structurally diverse metabolites are derived from a family of related enzymes called polyketide synthases (PKSs). A search of genomic sequence of Fusarium verticillioides, F. graminearum, F. oxysporum and Nectria haematococca (anamorph F. solani) identified a total of 58 PKS genes. To gain insight into how this gene family evolved and to guide future studies, we conducted a phylogenomic and functional domain analysis. The resulting genealogy suggested that Fusarium PKSs represent 34 different groups responsible for synthesis of different core metabolites. The analyses indicate that variation in the Fusarium PKS gene family is due to gene duplication and loss events as well as enzyme gain-of-function due to the acquisition of new domains or of loss-of-function due to nucleotide mutations. Transcriptional analysis indicate that the 16 F. verticillioides PKS genes are expressed under a range of conditions, further evidence that they are functional genes that confer the ability to produce secondary metabolites.

  7. Quantum spaces are modular

    NASA Astrophysics Data System (ADS)

    Freidel, Laurent; Leigh, Robert G.; Minic, Djordje

    2016-11-01

    At present, our notion of space is a classical concept. Taking the point of view that quantum theory is more fundamental than classical physics, and that space should be given a purely quantum definition, we revisit the notion of Euclidean space from the point of view of quantum mechanics. Since space appears in physics in the form of labels on relativistic fields or Schrödinger wave functionals, we propose to define Euclidean quantum space as a choice of polarization for the Heisenberg algebra of quantum theory. We show, following Mackey, that generically, such polarizations contain a fundamental length scale and that contrary to what is implied by the Schrödinger polarization, they possess topologically distinct spectra. These are the modular spaces. We show that they naturally come equipped with additional geometrical structures usually encountered in the context of string theory or generalized geometry. Moreover, we show how modular space reconciles the presence of a fundamental scale with translation and rotation invariance. We also discuss how the usual classical notion of space comes out as a form of thermodynamical limit of modular space while the Schrödinger space is a singular limit.

  8. [Modular enteral nutrition in pediatrics].

    PubMed

    Murillo Sanchís, S; Prenafeta Ferré, M T; Sempere Luque, M D

    1991-01-01

    Modular Enteral Nutrition may be a substitute for Parenteral Nutrition in children with different pathologies. Study of 4 children with different pathologies selected from a group of 40 admitted to the Maternal-Childrens Hospital "Valle de Hebrón" in Barcelona, who received modular enteral nutrition. They were monitored on a daily basis by the Dietician Service. Modular enteral nutrition consists of modules of proteins, peptides, lipids, glucids and mineral salts-vitamins. 1.--Craneo-encephalic traumatisms with loss of consciousness, Feeding with a combination of parenteral nutrition and modular enteral nutrition for 7 days. In view of the tolerance and good results of the modular enteral nutrition, the parenteral nutrition was suspended and modular enteral nutrition alone used up to a total of 43 days. 2.--55% burns with 36 days of hyperproteic modular enteral nutrition together with normal feeding. A more rapid recovery was achieved with an increase in total proteins and albumin. 3.--Persistent diarrhoea with 31 days of modular enteral nutrition, 5 days on parenteral nutrition alone and 8 days on combined parenteral nutrition and modular enteral nutrition. In view of the tolerance and good results of the modular enteral nutrition, the parenteral nutrition was suspended. 4.--Mucoviscidosis with a total of 19 days on modular enteral nutrition, 12 of which were exclusively on modular enteral nutrition and 7 as a night supplement to normal feeding. We administered proteic intakes of up to 20% of the total calorific intake and in concentrations of up to 1.2 calories/ml of the final preparation, always with a good tolerance. Modular enteral nutrition can and should be used as a substitute for parenteral nutrition in children with different pathologies, thus preventing the complications inherent in parenteral nutrition.

  9. Antibacterial polyketides from Bacillus amyloliquefaciens associated with edible red seaweed Laurenciae papillosa.

    PubMed

    Chakraborty, Kajal; Thilakan, Bini; Raola, Vamshi Krishna; Joy, Minju

    2017-03-01

    Heterotrophic Bacillus amyloliquefaciens associated with edible red seaweed, Laurenciae papillosa was used to isolate antibacterial polyketide compounds. Antibacterial activity studies integrated with the outcome obtained by polyketide synthetase (pks) coding genes established that seaweed-affiliated bacterial flora had a wide-ranging antibacterial activities and potential natural product diversity, which proved that the bacterium is valuable reservoir of novel bioactive metabolites. Bioactivity-guided isolation of 3-(octahydro-9-isopropyl-2H-benzo[h]chromen-4-yl)-2-methylpropyl benzoate and methyl 8-(2-(benzoyloxy)-ethyl)-hexahydro-4-((E)-pent-2-enyl)-2H-chromene-6-carboxylate of polyketide origin, with activity against human opportunistic food pathogenic microbes, have been isolated from the ethyl acetate extract of B. amyloliquefaciens. Structure-activity relationship analysis revealed that hydrophobic descriptor of the polyketide compounds significantly contribute towards its antibacterial activity. Seaweed-associated microorganisms were shown to represent a potential source of antimicrobial compounds for food and health benefits. The antibacterial polyketide compounds described in the present study may find potential applications in the food industry to reduce food-borne pathogens.

  10. A Turnstile Mechanism for the Controlled Growth of Biosynthetic Intermediates on Assembly Line Polyketide Synthases

    PubMed Central

    2016-01-01

    Vectorial polyketide biosynthesis on an assembly line polyketide synthase is the most distinctive property of this family of biological machines, while providing the key conceptual tool for the bioinformatic decoding of new antibiotic pathways. We now show that the action of the entire assembly line is synchronized by a previously unrecognized turnstile mechanism that prevents the ketosynthase domain of each module from being acylated by a new polyketide chain until the product of the prior catalytic cycle has been passed to the downstream module from the corresponding acyl carrier protein domain. The turnstile is closed by virtue of tight coupling to the signature decarboxylative condensation reaction catalyzed by the ketosynthase domain of each polyketide synthase module. Reopening of the turnstile is coupled to the eventual chain translocation step that vacates the module. At the maximal rate of substrate turnover, one would expect the chain release step to initiate a cascade of chain translocation events that sequentially migrate back upstream, thereby repriming each module and setting up the assembly line for the next round of polyketide chain elongation. PMID:26878060

  11. Expanding the chemical space of polyketides through structure-guided mutagenesis of Vitis vinifera stilbene synthase.

    PubMed

    Bhan, Namita; Cress, Brady F; Linhardt, Robert J; Koffas, Mattheos

    2015-08-01

    Several natural polyketides (PKs) have been associated with important pharmaceutical properties. Type III polyketide synthases (PKS) that generate aromatic PK polyketides have been studied extensively for their substrate promiscuity and product diversity. Stilbene synthase-like (STS) enzymes are unique in the type III PKS class as they possess a hydrogen bonding network, furnishing them with thioesterase-like properties, resulting in aldol condensation of the polyketide intermediates formed. Chalcone synthases (CHS) in contrast, lack this hydrogen-bonding network, resulting primarily in the Claisen condensation of the polyketide intermediates formed. We have attempted to expand the chemical space of this interesting class of compounds generated by creating structure-guided mutants of Vitis vinifera STS. Further, we have utilized a previously established workflow to quickly compare the wild-type reaction products to those generated by the mutants and identify novel PKs formed by using XCMS analysis of LC-MS and LC-MS/MS data. Based on this approach, we were able to generate 15 previously unreported PK molecules by exploring the substrate promiscuity of the wild-type enzyme and all mutants using unnatural substrates. These structures were specific to STSs and cannot be formed by their closely related CHS-like counterparts. Copyright © 2015 Elsevier B.V. and Société Française de Biochimie et Biologie Moléculaire (SFBBM). All rights reserved.

  12. LMDS Lightweight Modular Display System.

    DTIC Science & Technology

    1982-02-16

    LIGHTWEIGHT MODULAR DISPLAY SYSTEM %C AD Gomez SW Wolfe EW Davenport BD Calder 16 February 1982 * / DTrSJUL 22 3829 Approved for public release...375 4. TITLE (and Subtitle) S. TYPE OF REPORT & PERIOD COVERED Oct 77 to Jan 82 LMDS LIGHTWEIGHT MODULAR DISPLAY SYSTEM S. PERFORMING ORG. REPORT...Processing Power Distribution Modular Display Low Cost Tactical Display Tactical Tablet Lightweight Display General Purpose Dispiay Functional Modules Touch

  13. Characterization and transcriptional analysis of the gene cluster for coronafacic acid, the polyketide component of the phytotoxin coronatine.

    PubMed Central

    Liyanage, H; Palmer, D A; Ullrich, M; Bender, C L

    1995-01-01

    Coronafacic acid (CFA), the polyketide component of the phytotoxin coronatine (COR), is activated and coupled to coronamic acid via amide bond formation, a biosynthetic step presumably catalyzed by the CFA ligase (cfl) gene product. The COR biosynthetic gene cluster in Pseudomonas syringae pv. glycinea PG4180 is located within a 32-kb region of a 90-kb plasmid designated p4180A. In the present study, a cloned region of p4180A complemented all CFA- mutants spanning an 18.8-kb region of the COR biosynthetic cluster. The genetic evidence presented in this study indicates that cfl and the CFA biosynthetic gene cluster are encoded by a single transcript and that transcription of all of the genes in this operon is directed by the cfl promoter. The cfl promoter was localized to a 0.37-kb region upstream of the transcriptional start site by progressive subcloning in pRG960sd, a vector containing a promoterless glucuronidase gene. Transcription of the cfl/CFA operon was temperature sensitive and showed maximal glucuronidase activity at 18 degrees C. Furthermore, transcription of the cfl/CFA operon was dependent on the functional activity of a modified two-component regulatory system located within the COR biosynthetic gene cluster. Thermoregulation of the cfl/CFA operon and the coronamic acid biosynthetic gene cluster via the modified two-component regulatory system is discussed. PMID:8526495

  14. Expanding the fluorine chemistry of living systems using engineered polyketide synthase pathways

    PubMed Central

    Walker, Mark C.; Thuronyi, Benjamin W.; Charkoudian, Louise K.; Lowry, Brian; Khosla, Chaitan; Chang, Michelle C. Y.

    2014-01-01

    Organofluorines represent a rapidly expanding proportion of molecules used in pharmaceuticals, diagnostics, agrochemicals, and materials. Despite the prevalence of fluorine in synthetic compounds, the known biological scope is limited to a single pathway that produces fluoroacetate. Here, we demonstrate that this pathway can be exploited as a source of fluorinated building blocks for introduction of fluorine into natural product scaffolds. Specifically, we have constructed pathways involving two polyketide synthase systems and show that fluoroacetate can be used to incorporate fluorine into the polyketide backbone in vitro. We further show that fluorine can be introduced site-selectively and introduced into polyketide products in vivo. These results highlight the prospects for the production of complex fluorinated natural products using synthetic biology. PMID:24009388

  15. Trichophycin A, a Cytotoxic Linear Polyketide Isolated from a Trichodesmium thiebautii Bloom

    PubMed Central

    Bertin, Matthew J.; Wahome, Paul G.; Zimba, Paul V.; He, Haiyin; Moeller, Peter D. R.

    2017-01-01

    In an effort to isolate and characterize bioactive secondary metabolites from Trichodesmium thiebautii blooms, collected cyanobacteria biomass was subjected to bioassay-guided extraction and fractionation using the human colon cancer cell line HCT-116, resulting in the isolation and subsequent structure characterization of a linear polyketide trichophycin A (1). The planar structure of 1 was completed using 1D and 2D NMR spectroscopy and high-resolution electrospray ionization mass spectrometry (HRESIMS). Trichophycin A was moderately toxic against the murine neuroblastoma cell line Neuro-2A (EC50: 6.5 μM) and HCT-116 cells (EC50: 11.7 μM). Trichophycin A was significantly more cytotoxic than the previously isolated polyketides trichotoxin A and trichotoxin B. These cytotoxicity observations suggest that toxicity may be related to the polyol character of these polyketide compounds. PMID:28067831

  16. Bioinformatics tools for genome mining of polyketide and non-ribosomal peptides.

    PubMed

    Boddy, Christopher N

    2014-02-01

    Microbial natural products have played a key role in the development of clinical agents in nearly all therapeutic areas. Recent advances in genome sequencing have revealed that there is an incredible wealth of new polyketide and non-ribosomal peptide natural product diversity to be mined from genetic data. The diversity and complexity of polyketide and non-ribosomal peptide biosynthesis has required the development of unique bioinformatics tools to identify, annotate, and predict the structures of these natural products from their biosynthetic gene clusters. This review highlights and evaluates web-based bioinformatics tools currently available to the natural product community for genome mining to discover new polyketides and non-ribosomal peptides.

  17. Polyketides from the cultured lichen mycobiont of a Vietnamese Pyrenula sp.

    PubMed

    Le, Duy Hoang; Takenaka, Yukiko; Hamada, Nobuo; Mizushina, Yoshiyuki; Tanahashi, Takao

    2014-06-27

    A spore-derived mycobiont of a crustose Pyrenula sp. lichen collected in Vietnam was cultivated on a malt-yeast extract medium supplemented with 10% sucrose. Chemical investigation of the cultivated colonies led to the isolation of eight new alkylated decalin-type polyketides (1-8) along with three known compounds. The structures of these compounds were elucidated by spectroscopic and chemical means. This is the first instance of this type of polyketide being isolated from a cultured lichen mycobiont. The isolated polyketides 1 and 7 exhibited inhibitory activities against mammalian DNA polymerases α and β with IC50 values ranging from 8.1 to 19.5 μM. Compound 1 showed cytotoxic effects against the HCT116 human colon carcinoma cultured cell line with an IC50 value of 6.4 ± 0.7 μM.

  18. Modification of Rifamycin Polyketide Backbone Leads to Improved Drug Activity against Rifampicin-resistant Mycobacterium tuberculosis*

    PubMed Central

    Nigam, Aeshna; Almabruk, Khaled H.; Saxena, Anjali; Yang, Jongtae; Mukherjee, Udita; Kaur, Hardeep; Kohli, Puneet; Kumari, Rashmi; Singh, Priya; Zakharov, Lev N.; Singh, Yogendra; Mahmud, Taifo; Lal, Rup

    2014-01-01

    Rifamycin B, a product of Amycolatopsis mediterranei S699, is the precursor of clinically used antibiotics that are effective against tuberculosis, leprosy, and AIDS-related mycobacterial infections. However, prolonged usage of these antibiotics has resulted in the emergence of rifamycin-resistant strains of Mycobacterium tuberculosis. As part of our effort to generate better analogs of rifamycin, we substituted the acyltransferase domain of module 6 of rifamycin polyketide synthase with that of module 2 of rapamycin polyketide synthase. The resulting mutants (rifAT6::rapAT2) of A. mediterranei S699 produced new rifamycin analogs, 24-desmethylrifamycin B and 24-desmethylrifamycin SV, which contained modification in the polyketide backbone. 24-Desmethylrifamycin B was then converted to 24-desmethylrifamycin S, whose structure was confirmed by MS, NMR, and X-ray crystallography. Subsequently, 24-desmethylrifamycin S was converted to 24-desmethylrifampicin, which showed excellent antibacterial activity against several rifampicin-resistant M. tuberculosis strains. PMID:24923585

  19. Phenalenones: insight into the biosynthesis of polyketides from the marine alga-derived fungus Coniothyrium cereale.

    PubMed

    Nazir, Mamona; El Maddah, Fayrouz; Kehraus, Stefan; Egereva, Ekaterina; Piel, Jörn; Brachmann, Alexander O; König, Gabriele M

    2015-08-07

    The marine alga-derived fungus Coniothyrium cereale is a prolific producer of phenalenones. These polyketides were shown to possess antimicrobial effects and inhibitory activity towards the protease human leucocyte elastase (HLE). The current study focused on the biosynthesis of eight different structural types of phenalenones, comprising the natural products rousselianone A' (1), coniosclerodin (3), cereolactam (12), cereoaldomine (15), and trypethelone (16). Solid agar cultures of C. cereale were used to follow up the incorporation of [1-(13)C] labeled acetate into these metabolites. Taking the respective mechanisms of polyketide metabolism into account, the labeling pattern was interpreted, thus providing a hypothesis for the biosynthetic formation of the phenalenones. The polyketide skeleton of the phenanthrene-based compound cereolactam is proposed to be formed through degradation of a heptaketide by loss of two carbon atoms.

  20. The Use of ClusterMine360 for the Analysis of Polyketide and Nonribosomal Peptide Biosynthetic Pathways.

    PubMed

    Tremblay, Nicolas; Hill, Patrick; Conway, Kyle R; Boddy, Christopher N

    2016-01-01

    Polyketides and nonribosomal peptides constitute two large families of microbial natural products. Over the past 20 years a broad range of microbial polyketide and nonribosomal peptide biosynthetic pathways have been characterized leading to a surfeit of genetic data on polyketide and nonribosomal peptide biosynthesis. We developed the ClusterMine360 database, which stores the antiSMASH-based annotation of gene clusters in the NCBI database, linking the structure of the natural product to the biosynthetic gene cluster. This database is searchable and enables the user to access multiple sequence files for phylogenetic analysis of polyketide and nonribosomal peptide biosynthetic genes. Herein we describe how to add compound families and gene clusters to the database and search it using key words or structures to identify specific gene clusters. We also describe how to download multiple sequence files for specific catalytic domains from polyketide and nonribosomal peptide biosynthesis.

  1. Robotic hand with modular extensions

    DOEpatents

    Salisbury, Curt Michael; Quigley, Morgan

    2015-01-20

    A robotic device is described herein. The robotic device includes a frame that comprises a plurality of receiving regions that are configured to receive a respective plurality of modular robotic extensions. The modular robotic extensions are removably attachable to the frame at the respective receiving regions by way of respective mechanical fuses. Each mechanical fuse is configured to trip when a respective modular robotic extension experiences a predefined load condition, such that the respective modular robotic extension detaches from the frame when the load condition is met.

  2. Modular analysis of biological networks.

    PubMed

    Kaltenbach, Hans-Michael; Stelling, Jörg

    2012-01-01

    The analysis of complex biological networks has traditionally relied on decomposition into smaller, semi-autonomous units such as individual signaling pathways. With the increased scope of systems biology (models), rational approaches to modularization have become an important topic. With increasing acceptance of de facto modularity in biology, widely different definitions of what constitutes a module have sparked controversies. Here, we therefore review prominent classes of modular approaches based on formal network representations. Despite some promising research directions, several important theoretical challenges remain open on the way to formal, function-centered modular decompositions for dynamic biological networks.

  3. Biogenic antioxidative and anti-inflammatory aryl polyketides from the venerid bivalve clam Paphia malabarica.

    PubMed

    Joy, Minju; Chakraborty, Kajal

    2017-12-15

    Chemical investigation of ethyl acetate-methanol extract of the venerid bivalve clam Paphia malabarica led to isolation of three unprecedented aryl polyketide derivatives, characterized as (E)-12-(17-ethyl-tetrahydro-16-hydroxy-15-(methyl pentanoate)-14-oxo-2H-pyran-13-yl)-9-methyl-but-11-enyl benzoate (1), isobutyl-13-(6-(benzoyloxy)-10-methylpentyl)-tetrahydro-13-methyl-2H-pyran-17-carboxylate (2) and (13-(methoxycarbonyl)-11-((E)-18-ethylhexa-16,19-dienyl)-12-propyl-cyclohex-10-enyl)-methyl-3-hydroxy benzoate (3). The structures of the polyketides were assigned by extensive spectroscopic experiments. Compound 1 displayed comparatively greater 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical quenching potential (50% inhibitory concentration, IC50 ∼0.59mg/mL) than commercially available α-tocopherol (IC50 0.63mg/mL). Potential pro-inflammatory 5-lipoxygenase (5-LOX) inhibition potential (IC50 0.76-0.92mg/mL) of the polyketides in consonant with significantly greater anti-inflammatory selectivity indices (anti-cyclooxygense-1IC50/anti-cyclooxygense-2IC50>1) than non-steroidal anti-inflammatory agent ibuprofen (0.44) described the safety profile of the title compounds. Putative biosynthetic route by means of polyketide synthatase biocatalyzed pathways unambiguously established the structural assignments of the previously undescribed polyketide analogues. The potential of hitherto undescribed polyketides from P. malabarica as natural antioxidative and anti-inflammatory functional food ingredients was demonstrated in the present study. Copyright © 2017 Elsevier Ltd. All rights reserved.

  4. Discriminating the reaction types of plant type III polyketide synthases.

    PubMed

    Shimizu, Yugo; Ogata, Hiroyuki; Goto, Susumu

    2017-07-01

    Functional prediction of paralogs is challenging in bioinformatics because of rapid functional diversification after gene duplication events combined with parallel acquisitions of similar functions by different paralogs. Plant type III polyketide synthases (PKSs), producing various secondary metabolites, represent a paralogous family that has undergone gene duplication and functional alteration. Currently, there is no computational method available for the functional prediction of type III PKSs. We developed a plant type III PKS reaction predictor, pPAP, based on the recently proposed classification of type III PKSs. pPAP combines two kinds of similarity measures: one calculated by profile hidden Markov models (pHMMs) built from functionally and structurally important partial sequence regions, and the other based on mutual information between residue positions. pPAP targets PKSs acting on ring-type starter substrates, and classifies their functions into four reaction types. The pHMM approach discriminated two reaction types with high accuracy (97.5%, 39/40), but its accuracy decreased when discriminating three reaction types (87.8%, 43/49). When combined with a correlation-based approach, all 49 PKSs were correctly discriminated, and pPAP was still highly accurate (91.4%, 64/70) even after adding other reaction types. These results suggest pPAP, which is based on linear discriminant analyses of similarity measures, is effective for plant type III PKS function prediction. pPAP is freely available at ftp://ftp.genome.jp/pub/tools/ppap/. goto@kuicr.kyoto-u.ac.jp. Supplementary data are available at Bioinformatics online.

  5. Modular biometric system

    NASA Astrophysics Data System (ADS)

    Hsu, Charles; Viazanko, Michael; O'Looney, Jimmy; Szu, Harold

    2009-04-01

    Modularity Biometric System (MBS) is an approach to support AiTR of the cooperated and/or non-cooperated standoff biometric in an area persistent surveillance. Advanced active and passive EOIR and RF sensor suite is not considered here. Neither will we consider the ROC, PD vs. FAR, versus the standoff POT in this paper. Our goal is to catch the "most wanted (MW)" two dozens, separately furthermore ad hoc woman MW class from man MW class, given their archrivals sparse front face data basis, by means of various new instantaneous input called probing faces. We present an advanced algorithm: mini-Max classifier, a sparse sample realization of Cramer-Rao Fisher bound of the Maximum Likelihood classifier that minimize the dispersions among the same woman classes and maximize the separation among different man-woman classes, based on the simple feature space of MIT Petland eigen-faces. The original aspect consists of a modular structured design approach at the system-level with multi-level architectures, multiple computing paradigms, and adaptable/evolvable techniques to allow for achieving a scalable structure in terms of biometric algorithms, identification quality, sensors, database complexity, database integration, and component heterogenity. MBS consist of a number of biometric technologies including fingerprints, vein maps, voice and face recognitions with innovative DSP algorithm, and their hardware implementations such as using Field Programmable Gate arrays (FPGAs). Biometric technologies and the composed modularity biometric system are significant for governmental agencies, enterprises, banks and all other organizations to protect people or control access to critical resources.

  6. Modular Biometric Monitoring System

    NASA Technical Reports Server (NTRS)

    Chmiel, Alan J. (Inventor); Humphreys, Bradley T. (Inventor)

    2017-01-01

    A modular system for acquiring biometric data includes a plurality of data acquisition modules configured to sample biometric data from at least one respective input channel at a data acquisition rate. A representation of the sampled biometric data is stored in memory of each of the plurality of data acquisition modules. A central control system is in communication with each of the plurality of data acquisition modules through a bus. The central control system is configured to control communication of data, via the bus, with each of the plurality of data acquisition modules.

  7. Geometric Kac Moody modularity

    NASA Astrophysics Data System (ADS)

    Lynker, Monika; Schimmrigk, Rolf

    2006-05-01

    It is shown how the arithmetic structure of algebraic curves encoded in the Hasse-Weil L-function can be related to affine Kac-Moody algebras. This result is useful in relating the arithmetic geometry of Calabi-Yau varieties to the underlying exactly solvable theory. In the case of the genus three Fermat curve we identify the Hasse-Weil L-function with the Mellin transform of the twist of a number theoretic modular form derived from the string function of a non-twisted affine Lie algebra. The twist character is associated to the number field of quantum dimensions of the conformal field theory.

  8. Modular space station facilities.

    NASA Technical Reports Server (NTRS)

    Parker, P. J.

    1973-01-01

    The modular space station will operate as a general purpose laboratory (GPL). In addition, the space station will be able to support many attached or free-flying research and application modules that would be dedicated to specific projects like astronomy or earth observations. The GPL primary functions have been organized into functional laboratories including an electrical/electronics laboratory, a mechanical sciences laboratory, an experiment and test isolation laboratory, a hard data process facility, a data evaluation facility, an optical sciences laboratory, a biomedical and biosciences laboratory, and an experiment/secondary command and control center.

  9. Modular gear bearings

    NASA Technical Reports Server (NTRS)

    Vranish, John M. (Inventor)

    2009-01-01

    A gearing system using modular gear bearing components. Each component is composed of a core, one or more modules attached to the core and two or more fastening modules rigidly attaching the modules to the core. The modules, which are attached to the core, may consist of gears, rollers or gear bearing components. The core orientation affects the orientation of the modules attached to the core. This is achieved via the keying arrangement of the core and the component modules that attach to the core. Such an arrangement will also facilitate the phase tuning of gear modules with respect to the core and other gear modules attached to the core.

  10. Modularity of music processing.

    PubMed

    Peretz, Isabelle; Coltheart, Max

    2003-07-01

    The music faculty is not a monolithic entity that a person either has or does not. Rather, it comprises a set of neurally isolable processing components, each having the potential to be specialized for music. Here we propose a functional architecture for music processing that captures the typical properties of modular organization. The model rests essentially on the analysis of music-related deficits in neurologically impaired individuals, but provides useful guidelines for exploring the music faculty in normal people, using methods such as neuroimaging.

  11. Cervimycin A-D: a polyketide glycoside complex from a cave bacterium can defeat vancomycin resistance.

    PubMed

    Herold, Kerstin; Gollmick, Friedrich A; Groth, Ingrid; Roth, Martin; Menzel, Klaus-Dieter; Möllmann, Ute; Gräfe, Udo; Hertweck, Christian

    2005-09-19

    Cervimycins A-D are novel polyketide glycosides with significant activity against multi-drug-resistant staphylococci and vancomycin-resistant enterococci. They are produced by a strain of Streptomyces tendae, isolated from an ancient cave. The structures of the cervimycins were determined by performing extensive NMR and chemical degradation studies. All cervimycins have a common tetracyclic polyketide core that is substituted with unusual di- and tetrasaccharide chains, composed exclusively of trideoxysugars; however, they differ in the acetyl and carbamoyl ring substituent and in the highly unusual terminal methylmalonyl and dimethylmalonyl residues.

  12. Modular and Hierarchically Modular Organization of Brain Networks

    PubMed Central

    Meunier, David; Lambiotte, Renaud; Bullmore, Edward T.

    2010-01-01

    Brain networks are increasingly understood as one of a large class of information processing systems that share important organizational principles in common, including the property of a modular community structure. A module is topologically defined as a subset of highly inter-connected nodes which are relatively sparsely connected to nodes in other modules. In brain networks, topological modules are often made up of anatomically neighboring and/or functionally related cortical regions, and inter-modular connections tend to be relatively long distance. Moreover, brain networks and many other complex systems demonstrate the property of hierarchical modularity, or modularity on several topological scales: within each module there will be a set of sub-modules, and within each sub-module a set of sub-sub-modules, etc. There are several general advantages to modular and hierarchically modular network organization, including greater robustness, adaptivity, and evolvability of network function. In this context, we review some of the mathematical concepts available for quantitative analysis of (hierarchical) modularity in brain networks and we summarize some of the recent work investigating modularity of structural and functional brain networks derived from analysis of human neuroimaging data. PMID:21151783

  13. Modular antenna design study

    NASA Technical Reports Server (NTRS)

    Ribble, J. W.

    1981-01-01

    The mechanical design of a modular antenna concept was developed sufficiently to allow manufacture of a working demonstration model of a module, to predict mass properties, and to make performance estimates for antenna reflectors composed of these modules. The primary features of this concept are: (1) each module is an autonomous structural element which can be attached to adjacent modules through a three point connection; (2) the upper surface is a folding hexagonal truss plate mechanism which serves as the supporting structure for a reflective surface; and (3) the entire truss and surface can be folded into a cylindrical envelope in which all truss elements are essentially parallel. The kinematic studies and engineering demonstration model fully verified the deployment kinematics, stowing philosophy, and deployment sequencing for large antenna modules. It was established that such modules can be stowed in packages as small as 25 cm in diameter, using 1.27 cm diameter structural tubes. The development activity indicates that this deployable modular approach towards building large structures in space will support erection of 450 m apertures for operation up to 3 GHz with a single space shuttle flight.

  14. Modular reflector concept study

    NASA Technical Reports Server (NTRS)

    Vaughan, D. H.

    1981-01-01

    A study was conducted to evaluate the feasibility of space erecting a 100 meter paraboloidal radio frequency reflector by joining a number of individually deployed structural modules. Three module design concepts were considered: (1) the deployable cell module (DCM); (2) the modular paraboloidal erectable truss antenna (Mod-PETA); and (3) the modular erectable truss antenna (META). With the space shuttle (STS) as the launch system, the methodology of packaging and stowing in the orbiter, and of dispensing, deploying and joining, in orbit, were studied and the necessary support equipment identified. The structural performance of the completed reflectors was evaluated and their overall operational capability and feasibility were evaluated and compared. The potential of the three concepts to maintain stable shape in the space environment was determined. Their ability to operate at radio frequencies of 1 GHz and higher was assessed assuming the reflector surface to consist of a number of flat, hexagonal facets. A parametric study was performed to determine figure degradation as a function of reflector size, flat facet size, and f/D ratio.

  15. Modular radiochemistry synthesis system

    SciTech Connect

    Satyamurthy, Nagichettiar; Barrio, Jorge R.; Amarasekera, Bernard; Van Dam, R. Michael; Olma, Sebastian; Williams, Dirk; Eddings, Mark; Shen, Clifton Kwang-Fu

    2015-12-15

    A modular chemical production system includes multiple modules for performing a chemical reaction, particularly of radiochemical compounds, from a remote location. One embodiment comprises a reaction vessel including a moveable heat source with the position thereof relative to the reaction vessel being controllable from a remote position. Alternatively the heat source may be fixed in location and the reaction vial is moveable into and out of the heat source. The reaction vessel has one or more sealing plugs, the positioning of which in relationship to the reaction vessel is controllable from a remote position. Also the one or more reaction vessel sealing plugs can include one or more conduits there through for delivery of reactants, gases at atmospheric or an elevated pressure, inert gases, drawing a vacuum and removal of reaction end products to and from the reaction vial, the reaction vial with sealing plug in position being operable at elevated pressures. The modular chemical production system is assembled from modules which can each include operating condition sensors and controllers configured for monitoring and controlling the individual modules and the assembled system from a remote position. Other modules include, but are not limited to a Reagent Storage and Delivery Module, a Cartridge Purification Module, a Microwave Reaction Module, an External QC/Analysis/Purification Interface Module, an Aliquotting Module, an F-18 Drying Module, a Concentration Module, a Radiation Counting Module, and a Capillary Reactor Module.

  16. Modular radiochemistry synthesis system

    DOEpatents

    Satyamurthy, Nagichettiar; Barrio, Jorge R; Amarasekera, Bernard; Van Dam, R. Michael; Olma, Sebastian; Williams, Dirk; Eddings, Mark A; Shen, Clifton Kwang-Fu

    2015-02-10

    A modular chemical production system includes multiple modules for performing a chemical reaction, particularly of radiochemical compounds, from a remote location. One embodiment comprises a reaction vessel including a moveable heat source with the position thereof relative to the reaction vessel being controllable from a remote position. Alternatively the heat source may be fixed in location and the reaction vial is moveable into and out of the heat source. The reaction vessel has one or more sealing plugs, the positioning of which in relationship to the reaction vessel is controllable from a remote position. Also the one or more reaction vessel sealing plugs can include one or more conduits there through for delivery of reactants, gases at atmospheric or an elevated pressure, inert gases, drawing a vacuum and removal of reaction end products to and from the reaction vial, the reaction vial with sealing plug in position being operable at elevated pressures. The modular chemical production system is assembled from modules which can each include operating condition sensors and controllers configured for monitoring and controlling the individual modules and the assembled system from a remote position. Other modules include, but are not limited to a Reagent Storage and Delivery Module, a Cartridge Purification Module, a Microwave Reaction Module, an External QC/Analysis/Purification Interface Module, an Aliquotting Module, an F-18 Drying Module, a Concentration Module, a Radiation Counting Module, and a Capillary Reactor Module.

  17. Preheating after modular inflation

    NASA Astrophysics Data System (ADS)

    Barnaby, Neil; Bond, J. Richard; Huang, Zhiqi; Kofman, Lev

    2009-12-01

    We study (p)reheating in modular (closed string) inflationary scenarios, with a special emphasis on Kähler moduli/Roulette models. It is usually assumed that reheating in such models occurs through perturbative decays. However, we find that there are very strong non-perturbative preheating decay channels related to the particular shape of the inflaton potential (which is highly nonlinear and has a very steep minimum). Preheating after modular inflation, proceeding through a combination of tachyonic instability and broad-band parametric resonance, is perhaps the most violent example of preheating after inflation known in the literature. Further, we consider the subsequent transfer of energy to the standard model sector in scenarios where the standard model particles are confined to a D7-brane wrapping the inflationary blow-up cycle of the compactification manifold or, more interestingly, a non-inflationary blow-up cycle. We explicitly identify the decay channels of the inflaton in these two scenarios. We also consider the case where the inflationary cycle shrinks to the string scale at the end of inflation; here a field theoretical treatment of reheating is insufficient and one must turn instead to a stringy description. We estimate the decay rate of the inflaton and the reheat temperature for various scenarios.

  18. Modular Robotic Vehicle

    NASA Technical Reports Server (NTRS)

    Borroni-Bird, Christopher E. (Inventor); Vitale, Robert L. (Inventor); Lee, Chunhao J. (Inventor); Ambrose, Robert O. (Inventor); Bluethmann, William J. (Inventor); Junkin, Lucien Q. (Inventor); Lutz, Jonathan J. (Inventor); Guo, Raymond (Inventor); Lapp, Anthony Joseph (Inventor); Ridley, Justin S. (Inventor)

    2015-01-01

    A modular robotic vehicle includes a chassis, driver input devices, an energy storage system (ESS), a power electronics module (PEM), modular electronic assemblies (eModules) connected to the ESS via the PEM, one or more master controllers, and various embedded controllers. Each eModule includes a drive wheel containing a propulsion-braking module, and a housing containing propulsion and braking control assemblies with respective embedded propulsion and brake controllers, and a mounting bracket covering a steering control assembly with embedded steering controllers. The master controller, which is in communication with each eModule and with the driver input devices, communicates with and independently controls each eModule, by-wire, via the embedded controllers to establish a desired operating mode. Modes may include a two-wheel, four-wheel, diamond, and omni-directional steering modes as well as a park mode. A bumper may enable docking with another vehicle, with shared control over the eModules of the vehicles.

  19. Modular radiochemistry synthesis system

    SciTech Connect

    Satyamurthy, Nagichettiar; Barrio, Jorge R.; Amarasekera, Bernard; Van Dam, Michael R.; Olma, Sebastian; Williams, Dirk; Eddings, Mark; Shen, Clifton Kwang-Fu

    2016-11-01

    A modular chemical production system includes multiple modules for performing a chemical reaction, particularly of radiochemical compounds, from a remote location. One embodiment comprises a reaction vessel including a moveable heat source with the position thereof relative to the reaction vessel being controllable from a remote position. Alternatively the heat source may be fixed in location and the reaction vial is moveable into and out of the heat source. The reaction vessel has one or more sealing plugs, the positioning of which in relationship to the reaction vessel is controllable from a remote position. Also the one or more reaction vessel sealing plugs can include one or more conduits there through for delivery of reactants, gases at atmospheric or an elevated pressure, inert gases, drawing a vacuum and removal of reaction end products to and from the reaction vial, the reaction vial with sealing plug in position being operable at elevated pressures. The modular chemical production system is assembled from modules which can each include operating condition sensors and controllers configured for monitoring and controlling the individual modules and the assembled system from a remote position. Other modules include, but are not limited to a Reagent Storage and Delivery Module, a Cartridge Purification Module, a Microwave Reaction Module, an External QC/Analysis/Purification Interface Module, an Aliquotting Module, an F-18 Drying Module, a Concentration Module, a Radiation Counting Module, and a Capillary Reactor Module.

  20. Quantum modular forms, mock modular forms, and partial theta functions

    NASA Astrophysics Data System (ADS)

    Kimport, Susanna

    Defined by Zagier in 2010, quantum modular forms have been the subject of an explosion of recent research. Many of these results are aimed at discovering examples of these functions, which are defined on the rational numbers and have "nice" modularity properties. Though the subject is in its early stages, numerous results (including Zagier's original examples) show these objects naturally arising from many areas of mathematics as limits of other modular-like functions. One such family of examples is due to Folsom, Ono, and Rhoades, who connected these new objects to partial theta functions (introduced by Rogers in 1917) and mock modular forms (about which there is a rich theory, whose origins date back to Ramanujan in 1920). In this thesis, we build off of the work of Folsom, Ono, and Rhoades by providing an infinite family of quantum modular forms of arbitrary positive half-integral weight. Further, this family of quantum modular forms "glues" mock modular forms to partial theta functions and is constructed from a so-called "universal" mock theta function by extending a method of Eichler and Zagier (originally defined for holomorphic Jacobi forms) into a non-holomorphic setting. In addition to the infinite family, we explore the weight 1/2 and 3/2 functions in more depth. For both of these weights, we are able to explicitly write down the quantum modular form, as well as the corresponding "errors to modularity," which can be shown to be Mordell integrals of specific theta functions and, as a consequence, are real-analytic functions. Finally, we turn our attention to the partial theta functions associated with these low weight examples. Berndt and Kim provide asymptotic expansions for a certain class of partial theta functions as q approaches 1 radially within the unit disk. Here, we extend this work to not only obtain asymptotic expansions for this class of functions as q approaches any root of unity, but also for a certain class of derivatives of these functions

  1. Spacecraft Modularity for Serviceable Satellites

    NASA Technical Reports Server (NTRS)

    Rossetti, Dino; Keer, Beth; Panek, John; Ritter, Bob; Reed, Benjamin; Cepollina, Frank

    2015-01-01

    Spacecraft modularity has been a topic of interest at NASA since the 1970s, when the Multi-­-Mission Modular Spacecraft (MMS) was developed at the Goddard Space Flight Center. Since then, modular concepts have been employed for a variety of spacecraft and, as in the case of the Hubble Space Telescope (HST) and the International Space Station (ISS), have been critical to the success of on-­- orbit servicing. Modularity is even more important for future robotic servicing. Robotic satellite servicing technologies under development by NASA can extend mission life and reduce lifecycle cost and risk. These are optimized when the target spacecraft is designed for servicing, including advanced modularity. This paper will explore how spacecraft design, as demonstrated by the Reconfigurable Operational spacecraft for Science and Exploration (ROSE) spacecraft architecture, and servicing technologies can be developed in parallel to fully take advantage of the promise of both.

  2. Spacecraft Modularity for Serviceable Satellites

    NASA Technical Reports Server (NTRS)

    Reed, Benjamin B.; Rossetti, Dino; Keer, Beth; Panek, John; Cepollina, Frank; Ritter, Robert

    2015-01-01

    Spacecraft modularity has been a topic of interest at NASA since the 1970s, when the Multi-Mission Modular Spacecraft (MMS) was developed at the Goddard Space Flight Center. Since then, modular concepts have been employed for a variety of spacecraft and, as in the case of the Hubble Space Telescope (HST) and the International Space Station (ISS), have been critical to the success of on-orbit servicing. Modularity is even more important for future robotic servicing. Robotic satellite servicing technologies under development by NASA can extend mission life and reduce life-cycle cost and risk. These are optimized when the target spacecraft is designed for servicing, including advanced modularity. This paper will explore how spacecraft design, as demonstrated by the Reconfigurable Operational spacecraft for Science and Exploration (ROSE) spacecraft architecture, and servicing technologies can be developed in parallel to fully take advantage of the promise of both.

  3. A PLP-dependent polyketide chain releasing mechanism in the biosynthesis of mycotoxin fumonisins in Fusarium verticillioides.

    PubMed

    Gerber, Ryan; Lou, Lili; Du, Liangcheng

    2009-03-11

    Fumonisins are polyketide-derived mycotoxins produced by several plant pathogenic fungi. The toxins cause several fatal diseases in domestic animals and are associated with esophageal cancer and neural tube defects in humans. Fumonisins contain a highly reduced, acyclic 18-carbon chain, which is synthesized by an iterative polyketide synthase (PKS). This PKS does not contain a thioesterase or cyclase domain that is found in other PKSs for the release of the covalently linked polyketide chain. In this study, we expressed the acyl carrier protein (ACP) of FUM1 and in vitro loaded acyl chains to the ACP from acyl-CoA using a promiscuous 4'-phosphopantetheinyl transferase. We then expressed FUM8, which is homologous to 2-oxoamine synthase genes, in yeast and showed that the enzyme is able to offload the acyl chains from ACP. Products resulted from the decarboxylative condensation between l-alanine and acyl-S-ACP were detected by GC-MS. The enzyme activity was dependent on pyridoxal 5'-phosphate (PLP), and C18-S-ACP was the preferred substrate. The results revealed a novel polyketide chain-releasing mechanism, in which a PLP-dependent enzyme catalyzes the termination and offloading of the polyketide chain as well as the introduction of a new carbon-carbon bond and an amino group to the chain. The mechanism is fundamentally different from the thioesterase/cyclase-catalyzed polyketide chain release found in bacterial and other fungal polyketide biosyntheses.

  4. Modular Flooring System

    NASA Technical Reports Server (NTRS)

    Thate, Robert

    2012-01-01

    The modular flooring system (MFS) was developed to provide a portable, modular, durable carpeting solution for NASA fs Robotics Alliance Project fs (RAP) outreach efforts. It was also designed to improve and replace a modular flooring system that was too heavy for safe use and transportation. The MFS was developed for use as the flooring for various robotics competitions that RAP utilizes to meet its mission goals. One of these competitions, the FIRST Robotics Competition (FRC), currently uses two massive rolls of broadloom carpet for the foundation of the arena in which the robots are contained during the competition. The area of the arena is approximately 30 by 72 ft (approximately 9 by 22 m). This carpet is very cumbersome and requires large-capacity vehicles, and handling equipment and personnel to transport and deploy. The broadloom carpet sustains severe abuse from the robots during a regular three-day competition, and as a result, the carpet is not used again for competition. Similarly, broadloom carpets used for trade shows at convention centers around the world are typically discarded after only one use. This innovation provides a green solution to this wasteful practice. Each of the flooring modules in the previous system weighed 44 lb (.20 kg). The improvements in the overall design of the system reduce the weight of each module by approximately 22 lb (.10 kg) (50 %), and utilize an improved "module-to-module" connection method that is superior to the previous system. The MFS comprises 4-by-4-ft (.1.2-by- 1.2-m) carpet module assemblies that utilize commercially available carpet tiles that are bonded to a lightweight substrate. The substrate surface opposite from the carpeted surface has a module-to-module connecting interface that allows for the modules to be connected, one to the other, as the modules are constructed. This connection is hidden underneath the modules, creating a smooth, co-planar flooring surface. The modules are stacked and strapped

  5. Actinomycetes from the South China Sea sponges: isolation, diversity, and potential for aromatic polyketides discovery.

    PubMed

    Sun, Wei; Zhang, Fengli; He, Liming; Karthik, Loganathan; Li, Zhiyong

    2015-01-01

    Marine sponges often harbor dense and diverse microbial communities including actinobacteria. To date no comprehensive investigation has been performed on the culturable diversity of the actinomycetes associated with South China Sea sponges. Structurally novel aromatic polyketides were recently discovered from marine sponge-derived Streptomyces and Saccharopolyspora strains, suggesting that sponge-associated actinomycetes can serve as a new source of aromatic polyketides. In this study, a total of 77 actinomycete strains were isolated from 15 South China Sea sponge species. Phylogenetic characterization of the isolates based on 16S rRNA gene sequencing supported their assignment to 12 families and 20 genera, among which three rare genera (Marihabitans, Polymorphospora, and Streptomonospora) were isolated from marine sponges for the first time. Subsequently, β-ketoacyl synthase (KSα) gene was used as marker for evaluating the potential of the actinomycete strains to produce aromatic polyketides. As a result, KSα gene was detected in 35 isolates related to seven genera (Kocuria, Micromonospora, Nocardia, Nocardiopsis, Saccharopolyspora, Salinispora, and Streptomyces). Finally, 10 strains were selected for small-scale fermentation, and one angucycline compound was detected from the culture extract of Streptomyces anulatus strain S71. This study advanced our knowledge of the sponge-associated actinomycetes regarding their diversity and potential in producing aromatic polyketides.

  6. Phylogenetic Study of Polyketide Synthases and Nonribosomal Peptide Synthetases Involved in the Biosynthesis of Mycotoxins

    PubMed Central

    Gallo, Antonia; Ferrara, Massimo; Perrone, Giancarlo

    2013-01-01

    Polyketide synthase (PKSs) and nonribosomal peptide synthetase (NRPSs) are large multimodular enzymes involved in biosynthesis of polyketide and peptide toxins produced by fungi. Furthermore, hybrid enzymes, in which a reducing PKS region is fused to a single NRPS module, are also responsible of the synthesis of peptide-polyketide metabolites in fungi. The genes encoding for PKSs and NRPSs have been exposed to complex evolutionary mechanisms, which have determined the great number and diversity of metabolites. In this study, we considered the most important polyketide and peptide mycotoxins and, for the first time, a phylogenetic analysis of both PKSs and NRPSs involved in their biosynthesis was assessed using two domains for each enzyme: β-ketosynthase (KS) and acyl-transferase (AT) for PKSs; adenylation (A) and condensation (C) for NRPSs. The analysis of both KS and AT domains confirmed the differentiation of the three classes of highly, partially and non-reducing PKSs. Hybrid PKS-NRPSs involved in mycotoxins biosynthesis grouped together in the phylogenetic trees of all the domains analyzed. For most mycotoxins, the corresponding biosynthetic enzymes from distinct fungal species grouped together, except for PKS and NRPS involved in ochratoxin A biosynthesis, for which an unlike process of evolution could be hypothesized in different species. PMID:23604065

  7. A proteomic survey of nonribosomal peptide and polyketide biosynthesis in actinobacteria

    USDA-ARS?s Scientific Manuscript database

    Actinobacteria such as streptomycetes are renowned for their ability to produce bioactive natural products including nonribosomal peptides (NRPs) and polyketides (PKs). The advent of genome sequencing has revealed an even larger genetic repertoire for secondary metabolism with most of the small mole...

  8. Inversion of Extender Unit Selectivity in the Erythromycin Polyketide Synthase by Acyltransferase Domain Engineering.

    PubMed

    Koryakina, Irina; Kasey, Christian; McArthur, John B; Lowell, Andrew N; Chemler, Joseph A; Li, Shasha; Hansen, Douglas A; Sherman, David H; Williams, Gavin J

    2017-01-20

    Acyltransferase (AT) domains of polyketide synthases (PKSs) select extender units for incorporation into polyketides and dictate large portions of the structures of clinically relevant natural products. Accordingly, there is significant interest in engineering the substrate specificity of PKS ATs in order to site-selectively manipulate polyketide structure. However, previous attempts to engineer ATs have yielded mutant PKSs with relaxed extender unit specificity, rather than an inversion of selectivity from one substrate to another. Here, by directly screening the extender unit selectivity of mutants from active site saturation libraries of an AT from the prototypical PKS, 6-deoxyerythronolide B synthase, a set of single amino acid substitutions was discovered that dramatically impact the selectivity of the PKS with only modest reductions of product yields. One particular substitution (Tyr189Arg) inverted the selectivity of the wild-type PKS from its natural substrate toward a non-natural alkynyl-modified extender unit while maintaining more than twice the activity of the wild-type PKS with its natural substrate. The strategy and mutations described herein form a platform for combinatorial biosynthesis of site-selectively modified polyketide analogues that are modified with non-natural and non-native chemical functionality.

  9. c-di-GMP induction of Dictyostelium cell death requires the polyketide DIF-1.

    PubMed

    Song, Yu; Luciani, Marie-Françoise; Giusti, Corinne; Golstein, Pierre

    2015-02-15

    Cell death in the model organism Dictyostelium, as studied in monolayers in vitro, can be induced by the polyketide DIF-1 or by the cyclical dinucleotide c-di-GMP. c-di-GMP, a universal bacterial second messenger, can trigger innate immunity in bacterially infected animal cells and is involved in developmental cell death in Dictyostelium. We show here that c-di-GMP was not sufficient to induce cell death in Dictyostelium cell monolayers. Unexpectedly, it also required the DIF-1 polyketide. The latter could be exogenous, as revealed by a telling synergy between c-di-GMP and DIF-1. The required DIF-1 polyketide could also be endogenous, as shown by the inability of c-di-GMP to induce cell death in Dictyostelium HMX44A cells and DH1 cells upon pharmacological or genetic inhibition of DIF-1 biosynthesis. In these cases, c-di-GMP-induced cell death was rescued by complementation with exogenous DIF-1. Taken together, these results demonstrated that c-di-GMP could trigger cell death in Dictyostelium only in the presence of the DIF-1 polyketide or its metabolites. This identified another element of control to this cell death and perhaps also to c-di-GMP effects in other situations and organisms.

  10. Recent Advancements in the Biosynthetic Mechanisms for Polyketide-Derived Mycotoxins

    PubMed Central

    Huffman, Justin; Gerber, Ryan; Du, Liangcheng

    2010-01-01

    Polyketides are a large group of natural products produced by microorganisms and plants. They are biopolymers of acetate and other short carboxylates and are biosynthesized by multifunctional enzymes called polyketide synthases (PKSs). This review focuses on advances in molecular mechanisms of biosynthesis of four toxic polyketide, aflatoxins, fumonisins, ochratoxins, and zearalenone. These metabolites are structurally diverse and differ in their mechanisms of toxicity. However, they are all of concern in food safety and agriculture because of their toxic properties and their frequent accumulation in crops used for food and feed. The focus is on the recent advancements in the understanding of the molecular mechanisms for the biosynthesis of these mycotoxins. Several of the mycotoxin PKSs have been genetically and biochemically studied, while other PKSs remain to be investigated. Multiple post-PKS modifications are often required for the maturation of the mycotoxins. Many of these modification steps for aflatoxins and fumonisins are well established, while the post-PKS modifications for zearalenone and ochratoxins remain to be biochemically characterized. More efforts are needed in order to completely illustrate the biosynthetic mechanisms for this important group of polyketides. PMID:20578001

  11. Insights into natural products biosynthesis from analysis of 490 polyketide synthases from Fusarium.

    PubMed

    Brown, Daren W; Proctor, Robert H

    2016-04-01

    Species of the fungus Fusarium collectively cause disease on almost all crop plants and produce numerous natural products (NPs), including some of the mycotoxins of greatest concern to agriculture. Many Fusarium NPs are derived from polyketide synthases (PKSs), large multi-domain enzymes that catalyze sequential condensation of simple carboxylic acids to form polyketides. To gain insight into the biosynthesis of polyketide-derived NPs in Fusarium, we retrieved 488 PKS gene sequences from genome sequences of 31 species of the fungus. In addition to these apparently functional PKS genes, the genomes collectively included 81 pseudogenized PKS genes. Phylogenetic analysis resolved the PKS genes into 67 clades, and based on multiple lines of evidence, we propose that homologs in each clade are responsible for synthesis of a polyketide that is distinct from those synthesized by PKSs in other clades. The presence and absence of PKS genes among the species examined indicated marked differences in distribution of PKS homologs. Comparisons of Fusarium PKS genes and genes flanking them to those from other Ascomycetes provided evidence that Fusarium has the genetic potential to synthesize multiple NPs that are the same or similar to those reported in other fungi, but that have not yet been reported in Fusarium. The results also highlight ways in which such analyses can help guide identification of novel Fusarium NPs and differences in NP biosynthetic capabilities that exist among fungi.

  12. Altered expression of polyketide biosynthetic gene clusters in fumonisin-deficient mutants of Fusarium verticillioides

    USDA-ARS?s Scientific Manuscript database

    Fusarium verticillioides is a pathogen of maize and produces fumonisins, a group of polyketide derived secondary metabolites. Fumonisins cause diseases in animals, and they have been correlated epidemiologically with esophageal cancer and birth defects in humans. Fumonisin biosynthetic genes are c...

  13. Biochemical and Structural Characterization of the Tautomycetin Thioesterase: Analysis of a Stereoselective Polyketide Hydrolase

    SciTech Connect

    Scaglione, Jamie B.; Akey, David L.; Sullivan, Rachel; Kittendorf, Jeffrey D.; Rath, Christopher M.; Kim, Eung-Soo; Smith, Janet L.; Sherman, David H.

    2010-11-15

    A narrow tunnel: Biochemical and structural analysis of the tautomycetin thioesterase (TE) has provided the first high-resolution structure of a linear-chain-terminating TE in polyketide biosynthesis, showing the enzyme to be stereoselective with a constrained substrate chamber relative to macrolactone-forming thioesterases.

  14. Aromatic polyketide synthases from 127 Fusarium: pas de deux for chemical diversity

    USDA-ARS?s Scientific Manuscript database

    Fusarium species collectively cause disease on almost all crop plants and produce numerous natural products (NPs), including mycotoxins, of great concern. Many Fusarium NPs are derived from polyketide synthases (PKSs), large enzymes that catalyze the condensation of simple carboxylic acids. To gain ...

  15. Insights into natural products biosynthesis from analysis of 490 polyketide synthases from Fusarium

    USDA-ARS?s Scientific Manuscript database

    Species of the fungus Fusarium collectively cause disease on almost all crop plants and produce numerous natural products (NPs), including some of the mycotoxins of greatest concern to agriculture. Many Fusarium NPs are derived from polyketide synthases, large multi-domain enzymes that catalyze sequ...

  16. Identification of a Polyketide Synthase Coding Sequence Specific for Anatoxin-a-Producing Oscillatoria Cyanobacteria▿ †

    PubMed Central

    Cadel-Six, Sabrina; Iteman, Isabelle; Peyraud-Thomas, Caroline; Mann, Stéphane; Ploux, Olivier; Méjean, Annick

    2009-01-01

    We report the identification of a sequence from the genome of Oscillatoria sp. strain PCC 6506 coding for a polyketide synthase. Using 50 axenic cyanobacteria, we found this sequence only in the genomes of Oscillatoria strains producing anatoxin-a or homoanatoxin-a, indicating its likely involvement in the biosynthesis of these toxins. PMID:19447947

  17. A highly unusual polyketide synthase directs dawenol polyene biosynthesis in Stigmatella aurantiaca.

    PubMed

    Oßwald, Corina; Zaburannyi, Nestor; Burgard, Christian; Hoffmann, Thomas; Wenzel, Silke C; Müller, Rolf

    2014-12-10

    Enormous progress in the field of polyketide biosynthesis has led to the establishment of rules for general text book biosynthetic logic and consequently to the assumption that biosynthetic genes can be easily correlated with the corresponding natural products. However, non-textbook examples of polyketide assembly continue to be discovered suggesting the gene to product and product to gene predictions need improvement, especially as they are increasingly used in the post-genomic era. Here, we analyzed the genomic blueprint of a myxobacterial multi-producer of secondary metabolites, Stigmatella aurantiaca DW4/3-1, for its biosynthetic potential by genome-mining. In addition to the five polyketide synthase and/or nonribosomal peptide synthetase gene clusters of known function we identified a further 13 genomic regions exemplifying the enormous genetic potential for the production of additional chemical diversity by this strain. We show by gene inactivation and heterologous expression of the newly identified biosynthetic pathway for dawenol that the biosynthesis of this known polyene does not follow text book biosynthetic logic. Intriguingly, a genomic locus encoding an unusual polyketide synthase exhibiting similarity to gene loci involved in the formation of polyunsaturated fatty acids and secondary lipids was identified.

  18. Bipiperidine conjugates as soluble sugar surrogates in DNA-intercalating antiproliferative polyketides.

    PubMed

    Ueberschaar, Nico; Meyer, Florian; Dahse, Hans-Martin; Hertweck, Christian

    2016-04-07

    DNA-intercalating polyketide glycosides are important leads for cancer therapeutics, yet their use is often limited by their low solubility and challenging synthetic protocols. To overcome these limitations, we employed 1,4'-bipiperidine-1'-carbamate residues as sugar surrogates in daunorubicin and chartreusin, yielding water-soluble derivatives and prodrugs with dramatically improved antiproliferative activities.

  19. Starter unit specificity directs genome mining of polyketide synthase pathways in fungi

    USDA-ARS?s Scientific Manuscript database

    Search of the protein database with the aflatoxin pathway polyketide synthase (PKS) revealed putative PKSs in the pathogenic fungi Coccidioides immitis and Coccidioides posadasii that could require partnerships with a pair of fatty acid synthase (FAS) subunits for the biosynthesis of fatty acid-poly...

  20. Actinomycetes from the South China Sea sponges: isolation, diversity, and potential for aromatic polyketides discovery

    PubMed Central

    Sun, Wei; Zhang, Fengli; He, Liming; Karthik, Loganathan; Li, Zhiyong

    2015-01-01

    Marine sponges often harbor dense and diverse microbial communities including actinobacteria. To date no comprehensive investigation has been performed on the culturable diversity of the actinomycetes associated with South China Sea sponges. Structurally novel aromatic polyketides were recently discovered from marine sponge-derived Streptomyces and Saccharopolyspora strains, suggesting that sponge-associated actinomycetes can serve as a new source of aromatic polyketides. In this study, a total of 77 actinomycete strains were isolated from 15 South China Sea sponge species. Phylogenetic characterization of the isolates based on 16S rRNA gene sequencing supported their assignment to 12 families and 20 genera, among which three rare genera (Marihabitans, Polymorphospora, and Streptomonospora) were isolated from marine sponges for the first time. Subsequently, β-ketoacyl synthase (KSα) gene was used as marker for evaluating the potential of the actinomycete strains to produce aromatic polyketides. As a result, KSα gene was detected in 35 isolates related to seven genera (Kocuria, Micromonospora, Nocardia, Nocardiopsis, Saccharopolyspora, Salinispora, and Streptomyces). Finally, 10 strains were selected for small-scale fermentation, and one angucycline compound was detected from the culture extract of Streptomyces anulatus strain S71. This study advanced our knowledge of the sponge-associated actinomycetes regarding their diversity and potential in producing aromatic polyketides. PMID:26483773

  1. Modular arctic structures system

    SciTech Connect

    Reusswig, G. H.

    1984-12-04

    A modular and floatable offshore exploration and production platform system for use in shallow arctic waters is disclosed. A concrete base member is floated to the exploration or production site, and ballated into a predredged cavity. The cavity and base are sized to provide a stable horizontal base 30 feet below the mean water/ice plane. An exploration or production platform having a massive steel base is floated to the site and ballasted into position on the base. Together, the platform, base and ballast provide a massive gravity structure that is capable of resisting large ice and wave forces that impinge on the structure. The steel platform has a sloping hourglass profile to deflect horizontal ice loads vertically, and convert the horizontal load to a vertical tensile stress, which assists in breaking the ice as it advances toward the structure.

  2. Modular small hydro configuration

    NASA Astrophysics Data System (ADS)

    1981-09-01

    Smaller sites (those under 750 kilowatts) which previously were not attractive to develop using equipment intended for application at larger scale sites, were the focal point in the conception of a system which utilizes standard industrial components which are generally available within short procurement times. Such components were integrated into a development scheme for sites having 20 feet to 150 feet of head. The modular small hydro configuration maximizes the use of available components and minimizes modification of existing civil works. A key aspect of the development concept is the use of a vertical turbine multistage pump, used in the reverse mode as a hydraulic turbine. The configuration allows for automated operation and control of the hydroelectric facilities with sufficient flexibility for inclusion of potential hydroelectric sites into dispersed storage and generation (DSG) utility grid systems.

  3. Modular error embedding

    DOEpatents

    Sandford, II, Maxwell T.; Handel, Theodore G.; Ettinger, J. Mark

    1999-01-01

    A method of embedding auxiliary information into the digital representation of host data containing noise in the low-order bits. The method applies to digital data representing analog signals, for example digital images. The method reduces the error introduced by other methods that replace the low-order bits with auxiliary information. By a substantially reverse process, the embedded auxiliary data can be retrieved easily by an authorized user through use of a digital key. The modular error embedding method includes a process to permute the order in which the host data values are processed. The method doubles the amount of auxiliary information that can be added to host data values, in comparison with bit-replacement methods for high bit-rate coding. The invention preserves human perception of the meaning and content of the host data, permitting the addition of auxiliary data in the amount of 50% or greater of the original host data.

  4. Modular Optical PDV System

    SciTech Connect

    Araceli Rutkowski, David Esquibel

    2008-12-11

    A modular optical photon Doppler velocimetry (PDV) detector system has been developed by using readily available optical components with a 20-GHz Miteq optical detector into eight channels of single-wide modules integrated into a 3U rack unit (1U = 1.75 inches) with a common power supply. Optical fibers were precisely trimmed, welded, and timed within each unit. This system has been used to collect dynamic velocity data on various physics experiments. An optical power meter displays the laser input power to the module and optical power at the detector. An adjustable micro-electromechanical system (MEMS) optical attenuator is used to adjust the amount of unshifted light entering the detector. Front panel LEDs show the presence of power to the module. A fully loaded chassis with eight channels consumes 45 watts of power. Each chassis requires 1U spacing above and below for heat management. Modules can be easily replaced.

  5. Modular weapon control unit

    SciTech Connect

    Boccabella, M.F.; McGovney, G.N.

    1997-01-01

    The goal of the Modular Weapon Control Unit (MWCU) program was to design and develop a reconfigurable weapon controller (programmer/sequencer) that can be adapted to different weapon systems based on the particular requirements for that system. Programmers from previous systems are conceptually the same and perform similar tasks. Because of this commonality and the amount of re-engineering necessary with the advent of every new design, the idea of a modular, adaptable system has emerged. Also, the controller can be used in more than one application for a specific weapon system. Functionality has been divided into a Processor Module (PM) and an Input/Output Module (IOM). The PM will handle all operations that require calculations, memory, and timing. The IOM will handle interfaces to the rest of the system, input level shifting, output drive capability, and detection of interrupt conditions. Configuration flexibility is achieved in two ways. First, the operation of the PM is determined by a surface mount Read-Only Memory (ROM). Other surface-mount components can be added or neglected as necessary for functionality. Second, IOMs consist of configurable input buffers, configurable output drivers, and configurable interrupt generation. Further, these modules can be added singly or in groups to a Processor Module to achieve the required I/O configuration. The culmination of this LDRD was the building of both Processor Module and Input/Output Module. The MWCU was chosen as a test system to evaluate Low-Temperature Co-fired Ceramic (LTCC) technology, desirable for high component density and good thermal characteristics.

  6. Antimicrobial polyketide furanoterpenoids from seaweed-associated heterotrophic bacterium Bacillus subtilis MTCC 10403.

    PubMed

    Chakraborty, Kajal; Thilakan, Bini; Raola, Vamshi Krishna

    2017-10-01

    Brown seaweed Anthophycus longifolius (Turner) Kützing (family Sargassaceae) associated heterotrophic bacterium Bacillus subtilis MTCC 10403 was found to be a potent isolate with broad range of antibacterial activity against important perceptive food pathogens Vibrio parahaemolyticus, V. vulnificus, and Aeromonas hydrophila. This bacterium was positive for polyketide synthetase gene (KC589397), and therefore, was selected to bioprospect specialized metabolites bearing polyketide backbone. Bioactivity-guided chromatographic fractionation of the ethyl acetate extract of the seaweed-associated bacterium segregated four homologous polyketide furanoterpenoids with potential antibacterial activities against clinically important pathogens. The minimum inhibitory concentration (MIC) assay showed that the referral antibiotics tetracycline and ampicillin were active at 25 μg/mL against the test pathogens, whereas the previously undescribed (4E)-methyl 13-((16-(furan-2-yl) ethyl)-octahydro-7-hydroxy-4-((E)-23-methylbut-21-enyl)-2H-chromen-6-yl)-4-methylpent-4-enoate (compound 1) and methyl 3-(hexahydro-9-((E)-3-methylpent-1-enyl)-4H-furo[3,2-g]isochromen-6-yl) propanoate (compound 3) displayed antibacterial activities against the test pathogens at a lesser concentration (MIC < 7 μg/mL). The title compounds were characterized by comprehensive nuclear magnetic resonance and mass spectroscopic experiments. Polyketide synthase catalyzed putative biosynthetic mechanism additionally corroborated the structural ascriptions of the hitherto undescribed furanoterpenoids from seaweed-associated bacterial symbiont. The electronic and hydrophobic parameters appeared to hold a conspicuous part in directing the antibacterial properties of the compounds. Seaweed-associated B. subtilis MTCC 10403 demonstrated to represent a potential source of antimicrobial polyketides for pharmaceutical applications. Copyright © 2017 Elsevier Ltd. All rights reserved.

  7. Evolution of chemical diversity by coordinated gene swaps in type II polyketide gene clusters.

    PubMed

    Hillenmeyer, Maureen E; Vandova, Gergana A; Berlew, Erin E; Charkoudian, Louise K

    2015-11-10

    Natural product biosynthetic pathways generate molecules of enormous structural complexity and exquisitely tuned biological activities. Studies of natural products have led to the discovery of many pharmaceutical agents, particularly antibiotics. Attempts to harness the catalytic prowess of biosynthetic enzyme systems, for both compound discovery and engineering, have been limited by a poor understanding of the evolution of the underlying gene clusters. We developed an approach to study the evolution of biosynthetic genes on a cluster-wide scale, integrating pairwise gene coevolution information with large-scale phylogenetic analysis. We used this method to infer the evolution of type II polyketide gene clusters, tracing the path of evolution from the single ancestor to those gene clusters surviving today. We identified 10 key gene types in these clusters, most of which were swapped in from existing cellular processes and subsequently specialized. The ancestral type II polyketide gene cluster likely comprised a core set of five genes, a roster that expanded and contracted throughout evolution. A key C24 ancestor diversified into major classes of longer and shorter chain length systems, from which a C20 ancestor gave rise to the majority of characterized type II polyketide antibiotics. Our findings reveal that (i) type II polyketide structure is predictable from its gene roster, (ii) only certain gene combinations are compatible, and (iii) gene swaps were likely a key to evolution of chemical diversity. The lessons learned about how natural selection drives polyketide chemical innovation can be applied to the rational design and guided discovery of chemicals with desired structures and properties.

  8. The marine sponge-derived polyketide endoperoxide plakortide F acid mediates its antifungal activity by interfering with calcium homeostasis

    USDA-ARS?s Scientific Manuscript database

    Plakortide F acid (PFA) is a marine-derived polyketide endoperoxide exhibiting strong inhibitory activity against several clinically important fungal pathogens. In the present study, transcriptional profiling coupled with mutant and biochemical analyses were conducted using the model organism Sacch...

  9. Spacecraft Modularity for Serviceable Satellites

    NASA Technical Reports Server (NTRS)

    Rossetti, Dino; Keer, Beth; Panek, John; Reed, Benjamin; Cepollina, Frank; Ritter, Robert

    2015-01-01

    Satellite servicing has been a proven capability of NASA since the first servicing missions in the 1980s with astronauts on the space shuttle. This capability enabled the on-orbit assembly of the International Space Station (ISS) and saved the Hubble Space Telescope (HST) mission following the discovery of the flawed primary mirror. The effectiveness and scope of servicing opportunities, especially using robotic servicers, is a function of how cooperative a spacecraft is. In this paper, modularity will be presented as a critical design aspect for a spacecraft that is cooperative from a servicing perspective. Different features of modularity are discussed using examples from HST and the Multimission Modular Spacecraft (MMS) program from the 1980s and 1990s. The benefits of modularity will be presented including those directly related to servicing and those outside of servicing including reduced costs and increased flexibility. The new Reconfigurable Operational spacecraft for Science and Exploration (ROSE) concept is introduced as an affordable implementation of modularity that provides cost savings and flexibility. Key aspects of the ROSE architecture are discussed such as the module design and the distributed avionics architecture. The ROSE concept builds on the experience from MMS and due to its modularity, would be highly suitable as a future client for on-orbit servicing.

  10. The Modular Adaptive Ribosome.

    PubMed

    Yadav, Anupama; Radhakrishnan, Aparna; Panda, Anshuman; Singh, Amartya; Sinha, Himanshu; Bhanot, Gyan

    2016-01-01

    The ribosome is an ancient machine, performing the same function across organisms. Although functionally unitary, recent experiments suggest specialized roles for some ribosomal proteins. Our central thesis is that ribosomal proteins function in a modular fashion to decode genetic information in a context dependent manner. We show through large data analyses that although many ribosomal proteins are essential with consistent effect on growth in different conditions in yeast and similar expression across cell and tissue types in mice and humans, some ribosomal proteins are used in an environment specific manner. The latter set of variable ribosomal proteins further function in a coordinated manner forming modules, which are adapted to different environmental cues in different organisms. We show that these environment specific modules of ribosomal proteins in yeast have differential genetic interactions with other pathways and their 5'UTRs show differential signatures of selection in yeast strains, presumably to facilitate adaptation. Similarly, we show that in higher metazoans such as mice and humans, different modules of ribosomal proteins are expressed in different cell types and tissues. A clear example is nervous tissue that uses a ribosomal protein module distinct from the rest of the tissues in both mice and humans. Our results suggest a novel stratification of ribosomal proteins that could have played a role in adaptation, presumably to optimize translation for adaptation to diverse ecological niches and tissue microenvironments.

  11. The Modular Adaptive Ribosome

    PubMed Central

    Yadav, Anupama; Radhakrishnan, Aparna; Panda, Anshuman; Singh, Amartya; Sinha, Himanshu; Bhanot, Gyan

    2016-01-01

    The ribosome is an ancient machine, performing the same function across organisms. Although functionally unitary, recent experiments suggest specialized roles for some ribosomal proteins. Our central thesis is that ribosomal proteins function in a modular fashion to decode genetic information in a context dependent manner. We show through large data analyses that although many ribosomal proteins are essential with consistent effect on growth in different conditions in yeast and similar expression across cell and tissue types in mice and humans, some ribosomal proteins are used in an environment specific manner. The latter set of variable ribosomal proteins further function in a coordinated manner forming modules, which are adapted to different environmental cues in different organisms. We show that these environment specific modules of ribosomal proteins in yeast have differential genetic interactions with other pathways and their 5’UTRs show differential signatures of selection in yeast strains, presumably to facilitate adaptation. Similarly, we show that in higher metazoans such as mice and humans, different modules of ribosomal proteins are expressed in different cell types and tissues. A clear example is nervous tissue that uses a ribosomal protein module distinct from the rest of the tissues in both mice and humans. Our results suggest a novel stratification of ribosomal proteins that could have played a role in adaptation, presumably to optimize translation for adaptation to diverse ecological niches and tissue microenvironments. PMID:27812193

  12. Modular Approach to Spintronics.

    PubMed

    Camsari, Kerem Yunus; Ganguly, Samiran; Datta, Supriyo

    2015-06-11

    There has been enormous progress in the last two decades, effectively combining spintronics and magnetics into a powerful force that is shaping the field of memory devices. New materials and phenomena continue to be discovered at an impressive rate, providing an ever-increasing set of building blocks that could be exploited in designing transistor-like functional devices of the future. The objective of this paper is to provide a quantitative foundation for this building block approach, so that new discoveries can be integrated into functional device concepts, quickly analyzed and critically evaluated. Through careful benchmarking against available theory and experiment we establish a set of elemental modules representing diverse materials and phenomena. These elemental modules can be integrated seamlessly to model composite devices involving both spintronic and nanomagnetic phenomena. We envision the library of modules to evolve both by incorporating new modules and by improving existing modules as the field progresses. The primary contribution of this paper is to establish the ground rules or protocols for a modular approach that can build a lasting bridge between materials scientists and circuit designers in the field of spintronics and nanomagnetics.

  13. Modular Isotopic Thermoelectric Generator

    SciTech Connect

    Schock, Alfred

    1981-04-03

    Advanced RTG concepts utilizing improved thermoelectric materials and converter concepts are under study at Fairchild for DOE. The design described here is based on DOE's newly developed radioisotope heat source, and on an improved silicon-germanium material and a multicouple converter module under development at Syncal. Fairchild's assignment was to combine the above into an attractive power system for use in space, and to assess the specific power and other attributes of that design. The resultant design is highly modular, consisting of standard RTG slices, each producing ~24 watts at the desired output voltage of 28 volt. Thus, the design could be adapted to various space missions over a wide range of power levels, with little or no redesign. Each RTG slice consists of a 250-watt heat source module, eight multicouple thermoelectric modules, and standard sections of insulator, housing, radiator fins, and electrical circuit. The design makes it possible to check each thermoelectric module for electrical performance, thermal contact, leaktightness, and performance stability, after the generator is fully assembled; and to replace any deficient modules without disassembling the generator or perturbing the others. The RTG end sections provide the spring-loaded supports required to hold the free-standing heat source stack together during launch vibration. Details analysis indicates that the design offers a substantial improvement in specific power over the present generator of RTGs, using the same heat source modules. There are three copies in the file.

  14. Modular Approach to Spintronics

    PubMed Central

    Camsari, Kerem Yunus; Ganguly, Samiran; Datta, Supriyo

    2015-01-01

    There has been enormous progress in the last two decades, effectively combining spintronics and magnetics into a powerful force that is shaping the field of memory devices. New materials and phenomena continue to be discovered at an impressive rate, providing an ever-increasing set of building blocks that could be exploited in designing transistor-like functional devices of the future. The objective of this paper is to provide a quantitative foundation for this building block approach, so that new discoveries can be integrated into functional device concepts, quickly analyzed and critically evaluated. Through careful benchmarking against available theory and experiment we establish a set of elemental modules representing diverse materials and phenomena. These elemental modules can be integrated seamlessly to model composite devices involving both spintronic and nanomagnetic phenomena. We envision the library of modules to evolve both by incorporating new modules and by improving existing modules as the field progresses. The primary contribution of this paper is to establish the ground rules or protocols for a modular approach that can build a lasting bridge between materials scientists and circuit designers in the field of spintronics and nanomagnetics. PMID:26066079

  15. Engineered Biosynthesis of Plant Polyketides: Structure-Based and Precursor-Directed Approach

    NASA Astrophysics Data System (ADS)

    Abe, Ikuro

    Pentaketide chromone synthase (PCS) and octaketide synthase (OKS) are novel plant-specific type III polyketide synthases (PKSs) obtained from Aloe arborescens. Recombinant PCS expressed in Escherichia coli catalyzes iterative condensations of five molecules of malonyl-CoA to produce a pentaketide 5,7-dihydroxy-2-methylchromone, while recombinant OKS carries out sequential condensations of eight molecules of malonyl-CoA to yield octaketides SEK4 and SEK4b, the longest polyketides produced by the structurally simple type III PKS. The amino acid sequences of PCS and OKS are 91% identical, sharing 50-60% identity with those of other chalcone synthase (CHS) superfamily type III PKSs of plant origin. One of the most characteristic features is that the conserved active-site Thr197 of CHS (numbering in Medicago sativa CHS) is uniquely replaced with Met207 in PCS and with Gly207 in OKS, respectively. Site-directed mutagenesis and X-ray crystallographic analyses demonstrated that the chemically inert single residue lining the active-site cavity controls the polyketide chain length and the product specificity depending on the steric bulk of the side chain. On the basis of the crystal structures, an F80A/Y82A/M207G triple mutant of the pentaketide-producing PCS was constructed and shown to catalyze condensations of nine molecules of malonyl-CoA to produce an unnatural novel nonaketide naphthopyrone, whereas an N222G mutant of the octaketides-producing OKS yielded a decaketide benzophenone SEK15 from ten molecules of malonyl-CoA. On the other hand, the type III PKSs exhibited broad substrate specificities and catalytic potential. OKS accepted p-coumaroyl-CoA as a starter substrate to produce an unnatural novel C19 hexaketide stilbene and a C21 heptaketide chalcone. Remarkably, the C21 chalcone-forming activity was dramatically increased in the structure-guided OKS N222G mutant. In addition, OKS N222G mutant also yielded unnatural novel polyketides from phenylacetyl-CoA and

  16. Engineered biosynthesis of plant polyketides: structure-based and precursor-directed approach.

    PubMed

    Abe, Ikuro

    2010-01-01

    Pentaketide chromone synthase (PCS) and octaketide synthase (OKS) are novel plant-specific type III polyketide synthases (PKSs) obtained from Aloe arborescens. Recombinant PCS expressed in Escherichia coli catalyzes iterative condensations of five molecules of malonyl-CoA to produce a pentaketide 5,7-dihydroxy-2-methylchromone, while recombinant OKS carries out sequential condensations of eight molecules of malonyl-CoA to yield octaketides SEK4 and SEK4b, the longest polyketides produced by the structurally simple type III PKS. The amino acid sequences of PCS and OKS are 91% identical, sharing 50-60% identity with those of other chalcone synthase (CHS) superfamily type III PKSs of plant origin. One of the most characteristic features is that the conserved active-site Thr197 of CHS (numbering in Medicago sativa CHS) is uniquely replaced with Met207 in PCS and with Gly207 in OKS, respectively. Site-directed mutagenesis and X-ray crystallographic analyses demonstrated that the chemically inert single residue lining the active-site cavity controls the polyketide chain length and the product specificity depending on the steric bulk of the side chain. On the basis of the crystal structures, an F80A/Y82A/M207G triple mutant of the pentaketide-producing PCS was constructed and shown to catalyze condensations of nine molecules of malonyl-CoA to produce an unnatural novel nonaketide naphthopyrone, whereas an N222G mutant of the octaketides-producing OKS yielded a decaketide benzophenone SEK15 from ten molecules of malonyl-CoA. On the other hand, the type III PKSs exhibited broad substrate specificities and catalytic potential. OKS accepted p-coumaroyl-CoA as a starter substrate to produce an unnatural novel C19 hexaketide stilbene and a C21 heptaketide chalcone. Remarkably, the C21 chalcone-forming activity was dramatically increased in the structure-guided OKS N222G mutant. In addition, OKS N222G mutant also yielded unnatural novel polyketides from phenylacetyl-CoA and

  17. Two Polyketide Synthase-encoding Genes are Required for Biosynthesis of the Polyketide Virulence Factor, T-toxin, by Cochliobolus heterostrophus

    SciTech Connect

    Baker, Scott E.; Kroken, Scott; Inderbitzin, Patrik; Asvarak, Thipa; Li, Bi-Yu; Shi, Liang; Yoder, Olen C.; Turgeon, Barbara G.

    2006-03-01

    Cochliobolus heterostrophus race T, causal agent of Southern Corn Leaf Blight, requires T-toxin (a family of C35 – C49 polyketides) for high virulence on T-cytoplasm maize. Production of T-toxin is controlled by two unlinked loci, Tox1A and Tox1B, carried on 1.2 Mb of DNA not found in race O, a mildly virulent form of the fungus that does not produce T-toxin, or in any other Cochliobolus spp. or closely related fungus. PKS1, a polyketide synthase (PKS)-encoding gene at Tox1A and DEC1, a decarboxylase-encoding gene at Tox1B, are necessary for T-toxin production. Although there is evidence that additional genes are required for T-toxin production, efforts to clone them have been frustrated because the genes are located in highly repeated, A+T-rich DNA. To overcome this difficulty, Ligation specificity-based Expression Analysis Display (LEAD), a comparative AFLP/gel fractionation/capillary sequencing procedure was applied to cDNAs from a near isogenic pair of race T (Tox1+) and race O (Tox1-) strains. This led to discovery of PKS2, a second PKS-encoding gene that maps at Tox1A and is required for both T-toxin biosynthesis and high virulence to maize. Thus, the carbon chain of each T-toxin family member is likely assembled by action of two PKSs, which produce two polyketides, one of which may act as the starter unit for biosynthesis of the mature T-toxin molecule.

  18. Crystal Structure And Functional Analysis of Tetracenomycin Aro/Cyc: Implications for Cyclization Specificity of Aromatic Polyketides

    SciTech Connect

    Ames, B.D.; Korman, T.P.; Zhang, W.; Smith, P.; Vu, T.; Tang, Y.; Tsai, S.-C.

    2009-05-11

    Polyketides are a class of natural products with highly diverse chemical structures and pharmaceutical activities. Polyketide cyclization, promoted by the aromatase/cyclase (ARO/CYC), helps diversify aromatic polyketides. How the ARO/CYC promotes highly specific cyclization is not well understood because of the lack of a first-ring ARO/CYC structure. The 1.9 {angstrom} crystal structure of Tcm ARO/CYC reveals that the enzyme belongs to the Bet v1-like superfamily (or STAR domain family) with a helix-grip fold, and contains a highly conserved interior pocket. Docking, mutagenesis, and an in vivo assay show that the size, shape, and composition of the pocket are important to orient and specifically fold the polyketide chain for C9-C14 first-ring and C7-C16 second-ring cyclizations. Two pocket residues, R69 and Y35, were found to be essential for promoting first- and second-ring cyclization specificity. Different pocket residue mutations affected the polyketide product distribution. A mechanism is proposed based on the structure-mutation-docking results. These results strongly suggest that the regiospecific cyclizations of the first two rings and subsequent aromatizations take place in the interior pocket. The chemical insights gleaned from this work pave the foundation toward defining the molecular rules for the ARO/CYC cyclization specificity, whose rational control will be important for future endeavors in the engineered biosynthesis of novel anticancer and antibiotic aromatic polyketides.

  19. Structural and Biochemical Analyses of Regio- and Stereo-Specificities Observed in a Type II Polyketide Ketoreductase

    PubMed Central

    Javidpour, Pouya; Korman, Tyler Paz; Shakya, Gaurav; Tsai, Shiou-Chuan

    2011-01-01

    Type II polyketides include antibiotics such as tetracycline, and chemotherapeutics such as daunorubicin. Type II polyketides are biosynthesized by the type II polyketide synthase (PKS) that consists of 5 – 10 stand-alone domains. In many type II PKSs, the type II ketoreductase (KR) specifically reduce the C9-carbonyl group. How the type II KR achieves such a high regio-specificity, and the nature of stereo-specificity, are not well understood. Sequence alignment of KRs led to a hypothesis that a well-conserved 94-XGG-96 motif may be involved in controlling the stereochemistry. The stereo-specificity of single, double and triple mutant combinations of P94L, G95D and G96D were analyzed in vitro and in vivo for the actinorhodin KR (actKR). The P94L mutation is sufficient to change the stereospecificity of actKR. Binary and ternary crystal structures of both wild type and P94L actKR were solved. Together with assay results, docking simulations, and co-crystal structures, a model for stereochemical control is presented herein that elucidates how type II polyketides are introduced into the substrate pocket such that the C9-carbonyl can be reduced with high regio- and stereo-specificities. The molecular features of actKR important for regio- and stereo-specificities can potentially be applied to biosynthesize new polyketides via protein engineering that rationally controls polyketide ketoreduction. PMID:21506596

  20. Isolation and characterization of a reducing polyketide synthase gene from the lichen-forming fungus Usnea longissima.

    PubMed

    Wang, Yi; Kim, Jung A; Cheong, Yong Hwa; Joshi, Yogesh; Koh, Young Jin; Hur, Jae-Seoun

    2011-06-01

    The reducing polyketide synthases found in filamentous fungi are involved in the biosynthesis of many drugs and toxins. Lichens produce bioactive polyketides, but the roles of reducing polyketide synthases in lichens remain to be clearly elucidated. In this study, a reducing polyketide synthase gene (U1PKS3) was isolated and characterized from a cultured mycobiont of Usnea longissima. Complete sequence information regarding U1PKS3 (6,519 bp) was obtained by screening a fosmid genomic library. A U1PKS3 sequence analysis suggested that it contains features of a reducing fungal type I polyketide synthase with β-ketoacyl synthase (KS), acyltransferase (AT), dehydratase (DH), enoyl reductase (ER), ketoacyl reducatse (KR), and acyl carrier protein (ACP) domains. This domain structure was similar to the structure of ccRadsl, which is known to be involved in resorcylic acid lactone biosynthesis in Chaetomium chiversii. The results of phylogenetic analysis located U1PKS3 in the clade of reducing polyketide synthases. RT-PCR analysis results demonstrated that UIPKS3 had six intervening introns and that UIPKS3 expression was upregulated by glucose, sorbitol, inositol, and mannitol.

  1. Modular Stirling Radioisotope Generator

    NASA Technical Reports Server (NTRS)

    Schmitz, Paul C.; Mason, Lee S.; Schifer, Nicholas A.

    2015-01-01

    High efficiency radioisotope power generators will play an important role in future NASA space exploration missions. Stirling Radioisotope Generators (SRG) have been identified as a candidate generator technology capable of providing mission designers with an efficient, high specific power electrical generator. SRGs high conversion efficiency has the potential to extend the limited Pu-238 supply when compared with current Radioisotope Thermoelectric Generators (RTG). Due to budgetary constraints, the Advanced Stirling Radioisotope Generator (ASRG) was canceled in the fall of 2013. Over the past year a joint study by NASA and DOE called the Nuclear Power Assessment Study (NPAS) recommended that Stirling technologies continue to be explored. During the mission studies of the NPAS, spare SRGs were sometimes required to meet mission power system reliability requirements. This led to an additional mass penalty and increased isotope consumption levied on certain SRG-based missions. In an attempt to remove the spare power system, a new generator architecture is considered which could increase the reliability of a Stirling generator and provide a more fault-tolerant power system. This new generator called the Modular Stirling Radioisotope Generator (MSRG) employs multiple parallel Stirling convertor/controller strings, all of which share the heat from the General Purpose Heat Source (GPHS) modules. For this design, generators utilizing one to eight GPHS modules were analyzed, which provide about 50 to 450 watts DC to the spacecraft, respectively. Four Stirling convertors are arranged around each GPHS module resulting in from 4 to 32 Stirling/controller strings. The convertors are balanced either individually or in pairs, and are radiatively coupled to the GPHS modules. Heat is rejected through the housing/radiator which is similar in construction to the ASRG. Mass and power analysis for these systems indicate that specific power may be slightly lower than the ASRG and

  2. Modular Stirling Radioisotope Generator

    NASA Technical Reports Server (NTRS)

    Schmitz, Paul C.; Mason, Lee S.; Schifer, Nicholas A.

    2016-01-01

    High-efficiency radioisotope power generators will play an important role in future NASA space exploration missions. Stirling Radioisotope Generators (SRGs) have been identified as a candidate generator technology capable of providing mission designers with an efficient, high-specific-power electrical generator. SRGs high conversion efficiency has the potential to extend the limited Pu-238 supply when compared with current Radioisotope Thermoelectric Generators (RTGs). Due to budgetary constraints, the Advanced Stirling Radioisotope Generator (ASRG) was canceled in the fall of 2013. Over the past year a joint study by NASA and the Department of Energy (DOE) called the Nuclear Power Assessment Study (NPAS) recommended that Stirling technologies continue to be explored. During the mission studies of the NPAS, spare SRGs were sometimes required to meet mission power system reliability requirements. This led to an additional mass penalty and increased isotope consumption levied on certain SRG-based missions. In an attempt to remove the spare power system, a new generator architecture is considered, which could increase the reliability of a Stirling generator and provide a more fault-tolerant power system. This new generator called the Modular Stirling Radioisotope Generator (MSRG) employs multiple parallel Stirling convertor/controller strings, all of which share the heat from the General Purpose Heat Source (GPHS) modules. For this design, generators utilizing one to eight GPHS modules were analyzed, which provided about 50 to 450 W of direct current (DC) to the spacecraft, respectively. Four Stirling convertors are arranged around each GPHS module resulting in from 4 to 32 Stirling/controller strings. The convertors are balanced either individually or in pairs, and are radiatively coupled to the GPHS modules. Heat is rejected through the housing/radiator, which is similar in construction to the ASRG. Mass and power analysis for these systems indicate that specific

  3. Synthesis of (R)-mellein by a partially reducing iterative polyketide synthase.

    PubMed

    Sun, Huihua; Ho, Chun Loong; Ding, Feiqing; Soehano, Ishin; Liu, Xue-Wei; Liang, Zhao-Xun

    2012-07-25

    Mellein and the related 3,4-dihydroisocoumarins are a family of natural products with interesting biological properties. The mechanisms of dihydroisocoumarin biosynthesis remain largely speculative today. Here we report the synthesis of mellein by a partially reducing iterative polyketide synthase (PR-PKS) as a pentaketide product. Remarkably, despite the head-to-tail homology shared with several fungal and bacterial PR-PKSs, the mellein synthase exhibits a distinct keto reduction pattern in the synthesis of the pentaketide. We present evidence to show that the ketoreductase (KR) domain alone is able to recognize and differentiate the polyketide intermediates, which provides a mechanistic explanation for the programmed keto reduction in these PR-PKSs.

  4. Polyketide Quinones Are Alternate Intermediate Electron Carriers during Mycobacterial Respiration in Oxygen-Deficient Niches.

    PubMed

    Anand, Amitesh; Verma, Priyanka; Singh, Anil Kumar; Kaushik, Sandeep; Pandey, Rajesh; Shi, Ce; Kaur, Harneet; Chawla, Manbeena; Elechalawar, Chandra Kumar; Kumar, Dhirendra; Yang, Yong; Bhavesh, Neel S; Banerjee, Rajkumar; Dash, Debasis; Singh, Amit; Natarajan, Vivek T; Ojha, Anil K; Aldrich, Courtney C; Gokhale, Rajesh S

    2015-11-19

    Mycobacterium tuberculosis (Mtb) adaptation to hypoxia is considered crucial to its prolonged latent persistence in humans. Mtb lesions are known to contain physiologically heterogeneous microenvironments that bring about differential responses from bacteria. Here we exploit metabolic variability within biofilm cells to identify alternate respiratory polyketide quinones (PkQs) from both Mycobacterium smegmatis (Msmeg) and Mtb. PkQs are specifically expressed in biofilms and other oxygen-deficient niches to maintain cellular bioenergetics. Under such conditions, these metabolites function as mobile electron carriers in the respiratory electron transport chain. In the absence of PkQs, mycobacteria escape from the hypoxic core of biofilms and prefer oxygen-rich conditions. Unlike the ubiquitous isoprenoid pathway for the biosynthesis of respiratory quinones, PkQs are produced by type III polyketide synthases using fatty acyl-CoA precursors. The biosynthetic pathway is conserved in several other bacterial genomes, and our study reveals a redox-balancing chemicocellular process in microbial physiology.

  5. Biosynthesis of pyranonaphthoquinone polyketides reveals diverse strategies for enzymatic carbon-carbon bond formation.

    PubMed

    Metsä-Ketelä, Mikko; Oja, Terhi; Taguchi, Takaaki; Okamoto, Susumu; Ichinose, Koji

    2013-08-01

    Pyranonaphthoquinones synthesized by Streptomyces bacteria via type II polyketide pathways are aromatic compounds build around a common three-ring structure, which is composed of pyran, quinone and benzene rings. Over the years, actinorhodin in particular has served as a model compound for studying the biosynthesis of aromatic polyketides, while some of the other metabolites such as granaticin, medermycin, frenolicin and alnumycin A have enabled comparative studies that complement our understanding how these complex biological systems function and have evolved. In addition, despite the similarity of the aglycone units, pyranonaphthoquinones in effect display remarkable diversity in tailoring reactions, which include numerous enzymatic carbon-carbon bond forming reactions. This review focuses on the current status of molecular genetic, biochemical and structural investigations on this intriguing family of natural products.

  6. Broad Substrate Specificity of the Loading Didomain of the Lipomycin Polyketide Synthase

    SciTech Connect

    Yuzawa, S; Eng, CH; Katz, L; Keasling, JD

    2013-06-04

    LipPks1, a polyketide synthase subunit of the lipomycin synthase, is believed to catalyze the polyketide chain initiation reaction using isobutyryl-CoA as a substrate, followed by an elongation reaction with methylmalonyl-CoA to start the biosynthesis of antibiotic alpha-lipomycin in Streptomyces aureofaciens Tu117. Recombinant LipPks1, containing the thioesterase domain from the 6-deoxyerythronolide B synthase, was produced in Escherichia coli, and its substrate specificity was investigated in vitro. Surprisingly, several different acyl-CoAs, including isobutyryl-CoA, were accepted as the starter substrates, while no product was observed with acetyl-CoA. These results demonstrate the broad substrate specificity of LipPks1 and may be applied to producing new antibiotics.

  7. Tagging polyketides/non-ribosomal peptides with a clickable functionality and applications

    NASA Astrophysics Data System (ADS)

    Zhu, Xuejun; Zhang, Wenjun

    2015-02-01

    Bioorthogonal chemistry has recently emerged to be one of the most powerful tools in drug discovery and chemical biology. The exploration of it has successfully advanced the field of natural product research. In this Perspective, we survey current strategies for the installation of chemical handles into the molecular scaffolds of several major classes of natural products, including polyketides, non-ribosomal peptides, and their hybrids. By tagging these natural products with chemical handles and coupling them with subsequent bioorthogonal reactions, researchers have visualized and studied the mode of action of natural products, as well as synthesized derivatives with better pharmaceutical properties. We conclude this Perspective by considering two questions: Is there a general way to synthesize tagged polyketides/non-ribosomal peptides? Does natural product labeling have a broader impact in the field of natural product research beyond current known applications?

  8. Structural Analysis of Protein-Protein Interactions in Type I Polyketide Synthases

    PubMed Central

    Xu, Wei; Qiao, Kangjian; Tang, Yi

    2013-01-01

    Polyketide synthases (PKSs) are responsible for synthesizing a myriad of natural products with agricultural, medicinal relevance. The PKSs consist of multiple functional domains of which each can catalyze a specified chemical reaction leading to the synthesis of polyketides. Biochemical studies showed that protein-substrate and protein-protein interactions play crucial roles in these complex regio-/stereo- selective biochemical processes. Recent developments on X-ray crystallography and protein NMR techniques have allowed us to understand the biosynthetic mechanism of these enzymes from their structures. These structural studies have facilitated the elucidation of sequence-function relationship of PKSs and will ultimately contribute to the prediction of product structure. This review will focus on the current knowledge of type I PKS structures and the protein-protein interactions in this system. PMID:23249187

  9. The first plant type III polyketide synthase that catalyzes formation of aromatic heptaketide.

    PubMed

    Abe, Ikuro; Utsumi, Yoriko; Oguro, Satoshi; Noguchi, Hiroshi

    2004-03-26

    A cDNA encoding a novel plant type III polyketide synthase (PKS) was cloned from rhubarb (Rheum palmatum). A recombinant enzyme expressed in Escherichia coli accepted acetyl-CoA as a starter, carried out six successive condensations with malonyl-CoA and subsequent cyclization to yield an aromatic heptaketide, aloesone. The enzyme shares 60% amino acid sequence identity with chalcone synthases (CHSs), and maintains almost identical CoA binding site and catalytic residues conserved in the CHS superfamily enzymes. Further, homology modeling predicted that the 43-kDa protein has the same overall fold as CHS. This provides new insights into the catalytic functions of type III PKSs, and suggests further involvement in the biosynthesis of plant polyketides.

  10. Allelopathic Polyketides from an Endolichenic Fungus Myxotrichum SP. by Using OSMAC Strategy

    PubMed Central

    Yuan, Chao; Guo, Yu-Hua; Wang, Hai-Ying; Ma, Xiao-Jun; Jiang, Tao; Zhao, Jun-Ling; Zou, Zhong-Mei; Ding, Gang

    2016-01-01

    Three new polyketides myxotritones A-C (2–4), together with a new natural product 7,8-dihydro-7R,8S-dihydroxy-3,7-dimethyl-2-benzopyran-6-one (1) were obtained from the endolichenic fungus Myxotrichum sp. by using OMSAC (One Strain, Many Compounds) method. The planar structures of these new compounds were determined by NMR experiment and HRESIMS data, and the absolute configuration of 1 was established by X-ray diffraction, and the stereochemistry of the new compounds 2-4 were determined by same biosynthesis origin, and similar CD spectra with 1. Allelopathic test showed that compound 4 significantly retarded root elongation of Arabidopsis thaliana seed, indicating that this fungus might contribute to the defense of its host lichen. From the view of biosynthetic pathway, all four compounds 1-4 might be originated from Non-Reduced Polyketide synthase (NR-PKS). PMID:26839041

  11. Polyketide decarboxylative chain termination preceded by o-sulfonation in curacin a biosynthesis.

    PubMed

    Gu, Liangcai; Wang, Bo; Kulkarni, Amol; Gehret, Jennifer J; Lloyd, Kayla R; Gerwick, Lena; Gerwick, William H; Wipf, Peter; Håkansson, Kristina; Smith, Janet L; Sherman, David H

    2009-11-11

    Biosynthetic innovation in natural product systems is driven by the recruitment of new genes and enzymes into these complex pathways. Here, an unprecedented decarboxylative chain termination mechanism is described for the polyketide synthase of curacin A, an anticancer lead compound isolated from the marine cyanobacterium Lyngbya majuscula. The unusual chain termination module containing adjacent sulfotransferase (ST) and thioesterase (TE) catalytic domains embedded in CurM was biochemically characterized. The TE was proved to catalyze a hydrolytic chain release of the polyketide chain elongation intermediate. Moreover, a selective ST-mediated sulfonation of the (R)-beta-hydroxyl group was found to precede TE-mediated hydrolysis, triggering a successive decarboxylative elimination and resulting in the formation of a rare terminal olefin in the final metabolite.

  12. Phylogenomic and Domain Analysis of Iterative Polyketide Synthases in Aspergillus Species

    PubMed Central

    Lin, Shu-Hsi; Yoshimoto, Miwa; Lyu, Ping-Chiang; Tang, Chuan-Yi; Arita, Masanori

    2012-01-01

    Aspergillus species are industrially and agriculturally important as fermentors and as producers of various secondary metabolites. Among them, fungal polyketides such as lovastatin and melanin are considered a gold mine for bioactive compounds. We used a phylogenomic approach to investigate the distribution of iterative polyketide synthases (PKS) in eight sequenced Aspergilli and classified over 250 fungal genes. Their genealogy by the conserved ketosynthase (KS) domain revealed three large groups of nonreducing PKS, one group inside bacterial PKS, and more than 9 small groups of reducing PKS. Polyphyly of nonribosomal peptide synthase (NRPS)-PKS genes raised questions regarding the recruitment of the elegant conjugation machinery. High rates of gene duplication and divergence were frequent. All data are accessible through our web database at http://metabolomics.jp/wiki/Category:PK. PMID:22844193

  13. Discovery of clostrubin, an exceptional polyphenolic polyketide antibiotic from a strictly anaerobic bacterium.

    PubMed

    Pidot, Sacha; Ishida, Keishi; Cyrulies, Michael; Hertweck, Christian

    2014-07-21

    Genome mining of the strictly anaerobic bacterium Clostridium beijerinckii, an industrial producer of solvents, revealed the presence of several cryptic gene clusters for secondary metabolite biosynthesis. To unearth its metabolic potential, a C. beijerinckii strain was cultured under various conditions, which led to the discovery of a deep purple pigment. This novel metabolite, named clostrubin (1), was isolated and its structure was fully elucidated. The pentacyclic polyphenol features a benzo[a]tetraphene ring topology that is unprecedented for natural products. Stable-isotope labeling experiments showed that 1 is an aromatic polyketide that folds in a noncanonical manner to form the unusual perifused ring system. In addition to being the first reported polyketide from an anaerobic bacterium, 1 is a potent antibiotic with pronounced activity against various pathogenic bacteria, such as MRSA, VRE, and mycobacteria, with minimum inhibitory concentrations (MIC) of 0.12-0.97 μM.

  14. Designed biosynthesis of 36-methyl-FK506 by polyketide precursor pathway engineering

    PubMed Central

    Lechner, Anna; Wilson, Micheal C.; Ban, Yeon Hee; Hwang, Jae-yeon; Yoon, Yeo Joon; Moore, Bradley S.

    2012-01-01

    The polyketide synthase (PKS) biosynthetic code has recently expanded to include a newly recognized group of extender unit substrates derived from α,β-unsaturated acyl-CoA molecules that deliver diverse side chain chemistry to polyketide backbones. Herein we report the identification of a three-gene operon responsible for the biosynthesis of the PKS building block isobutyrylmalonyl-CoA associated with the macrolide ansalactam A from the marine bacterium Streptomyces sp. CNH189. Using a synthetic biology approach, we engineered the production of unnatural 36-methyl-FK506 in Streptomyces sp. KCTC 11604BP by incorporating the branched extender unit into FK506 biosynthesis in place of its natural C-21 allyl side chain, which has been shown to be critical for FK506’s potent immunosuppressant and neurite outgrowth activities. PMID:23654255

  15. Tafuketide, a phylogeny-guided discovery of a new polyketide from Talaromyces funiculosus Salicorn 58.

    PubMed

    Guo, Jia; Ran, Huomiao; Zeng, Jie; Liu, Dong; Xin, Zhihong

    2016-06-01

    A phylogeny-guided approach was applied to screen endophytic fungi containing type I polyketide synthase (PKS I) biosynthetic gene sequences and aimed to correlate genotype to chemotype for the discovery of novel bioactive polyketides. Salicorn 58, which was identified as Talaromyces funiculosus based on its internal transcribed spacer (ITS) and ribosomal large-subunit (LSU) DNA sequences, showed significant target bands. A chemical investigation of the culture of Salicorn 58 was allowed for the isolation of a new polyketide, Talafun (1), and a new natural product, N-(2'-hydroxy-3'-octadecenoyl)-9-methyl-4,8-sphingadienin (2), together with six known compounds, including chrodrimanin A (3), chrodrimanin B (4), N-(4-hydroxy-2-methoxyphenyl) acetamide (5), butyl β-glucose (6), 3β,15β-dihydroxyl-(22E, 24R)-ergosta-5,8(14),22-trien-7-dione (7), and (3β,5a,8a,22E)-5,8-epidioxyergosta-6,22-dien-3-ol (8). Their chemical structures were elucidated by extensive spectroscopic analysis and electro circular dichroism (ECD) spectrum calculations. Antioxidant experiments revealed that compound 5 showed strong ABTS(+) radical scavenging activity with an IC50 value of 11.43 ± 1.61 μM and potent ferric reducing activity (FRAP assay) with FRAP value of 187.52 ± 2.97. Antimicrobial assays revealed that compounds 1 and 4 showed high levels of selectivity toward Escherichia coli with MIC values of 18 ± 0.40 and 43 ± 0.52 μM, respectively. Compounds 2 and 3 exhibited broad-spectrum antimicrobial activity against Staphylococcus aureus, Mycobacterium smegmatis, Micrococcus tetragenus, Mycobacterium phlei, and E. coli, respectively. The results from the current research highlight the advantage of phylogeny-guided pipeline for the screening of new polyketides from endophytic fungi containing PKS I genes.

  16. Classification, prediction, and verification of the regioselectivity of fungal polyketide synthase product template domains.

    PubMed

    Li, Yanran; Image, Image Image; Xu, Wei; Image, Image; Tang, Yi; Image, Image

    2010-07-23

    The fungal iterative nonreducing polyketide synthases (NRPKSs) synthesize aromatic polyketides, many of which have important biological activities. The product template domains (PT) embedded in the multidomain NRPKSs mediate the regioselective cyclization of the highly reactive polyketide backbones and dictate the final structures of the products. Understanding the sequence-activity relationships of different PT domains is therefore an important step toward the prediction of polyketide structures from NRPKS sequences and can enable the genome mining of hundreds of cryptic NRPKSs uncovered via genome sequencing. In this work, we first performed phylogenetic analysis of PT domains from NRPKSs of known functions and showed that the PT domains can be classified into five groups, with each group corresponding to a unique product size or cyclization regioselectivity. Group V contains the formerly unverified PT domains that were identified as C6-C11 aldol cyclases. The regioselectivity of PTs from this group were verified by product-based assays using the PT domain excised from the asperthecin AptA NRPKS. When combined with dissociated PKS4 minimal PKS, or replaced the endogenous PKS4 C2-C7 PT domain in a hybrid NRPKS, AptA-PT directed the C6-C11 cyclization of the nonaketide backbone to yield a tetracyclic pyranoanthraquinone 4. Extensive NMR analysis verified that the backbone of 4 was indeed cyclized with the expected regioselectivity. The PT phylogenetic analysis was then expanded to include approximately 100 PT sequences from unverified NRPKSs. Using the assays developed for AptA-PT, the regioselectivities of additional PT domains were investigated and matched to those predicted by the phylogenetic classifications.

  17. First biosynthetic evidence on the phenyl-containing polyketides of the marine mollusc Scaphander lignarius.

    PubMed

    Cutignano, Adele; Avila, Conxita; Domenech-Coll, Anna; d'Ippolito, Giuliana; Cimino, Guido; Fontana, Angelo

    2008-07-17

    The biosynthesis of lignarenones 1 and 2, the major polyketides of the Mediterranean mollusc Scaphander lignarius is described. The process is primed by benzoic acid and requires acetate and propionate as extender units. The labeling pattern suggests PKS-like synthesis of an unusual E,Z,E-triene chain and origin of the benzoate unit from phenylalanine. 13C-13C NMR COSY has been used to establish the labeling positions due to incorporation of 13C2-acetate.

  18. Bacterial Fatty Acid Synthesis and its Relationships with Polyketide Synthetic Pathways

    PubMed Central

    Cronan, John E.; Thomas, Jacob

    2014-01-01

    This review presents the most thoroughly studied bacterial fatty acid synthetic pathway, that of Escherichia coli and then discusses the exceptions to the E. coli pathway present in other bacteria. The known interrelationships between the fatty acid and polyketide synthetic pathways are also assessed, mainly in the Streptomyces group of bacteria. Finally, we present a compendium of methods for analysis of bacterial fatty acid synthetic pathways. PMID:19362649

  19. Genome Mining of a Prenylated and Immunosuppressive Polyketide from Pathogenic Fungi

    PubMed Central

    Chooi, Yit-Heng; Fang, Jinxu; Liu, Hong; Filler, Scott G.; Wang, Pin; Tang, Yi

    2013-01-01

    Activation of the polycyclic polyketide prenyltransferases (pcPTase)-containing silent clusters in Aspergillus fumigatus and Neosartorya fischeri led to isolation of a new metabolite neosartoricin (3). The structure of 3 was solved by X-ray crystallography and NMR to be a prenylated anthracenone. 3 Exhibits T-cell antiproliferative activity with an IC50 of 3 μM, suggestive of a physiological role as an immunosuppressive agent. PMID:23368997

  20. Tanzawaic acids I–L: Four new polyketides from Penicillium sp. IBWF104-06

    PubMed Central

    Sandjo, Louis P; Thines, Eckhard

    2014-01-01

    Summary Four new polyketides have been identified in culture filtrates of the fungal strain Penicillium sp. IBWF104-06 isolated from a soil sample. They are structurally based on the same trans-decalinpentanoic acid skeleton as tanzawaic acids A–H. One of the new compounds was found to inhibit the conidial germination in the rice blast fungus Magnaporthe oryzae at concentrations of 25 μg/mL. PMID:24605144

  1. Heterologous expression in Saccharopolyspora erythraea of a pentaketide synthase derived from the spinosyn polyketide synthase.

    PubMed

    Martin, Christine J; Timoney, Máire C; Sheridan, Rose M; Kendrew, Steven G; Wilkinson, Barrie; Staunton, James C; Leadlay, Peter F

    2003-12-07

    A truncated version of the spinosyn polyketide synthase comprising the loading module and the first four extension modules fused to the erythromycin thioesterase domain was expressed in Saccharopolyspora erythraea. A novel pentaketide lactone product was isolated, identifying cryptic steps of spinosyn biosynthesis and indicating the potential of this approach for the biosynthetic engineering of spinosyn analogues. A pathway for the formation of the tetracyclic spinosyn aglycone is proposed.

  2. Modular Power Standard for Space Explorations Missions

    NASA Technical Reports Server (NTRS)

    Oeftering, Richard C.; Gardner, Brent G.

    2016-01-01

    Future human space exploration will most likely be composed of assemblies of multiple modular spacecraft elements with interconnected electrical power systems. An electrical system composed of a standardized set modular building blocks provides significant development, integration, and operational cost advantages. The modular approach can also provide the flexibility to configure power systems to meet the mission needs. A primary goal of the Advanced Exploration Systems (AES) Modular Power System (AMPS) project is to establish a Modular Power Standard that is needed to realize these benefits. This paper is intended to give the space exploration community a "first look" at the evolving Modular Power Standard and invite their comments and technical contributions.

  3. Eigenvalue Spectra of Modular Networks

    NASA Astrophysics Data System (ADS)

    Peixoto, Tiago P.

    2013-08-01

    A large variety of dynamical processes that take place on networks can be expressed in terms of the spectral properties of some linear operator which reflects how the dynamical rules depend on the network topology. Often, such spectral features are theoretically obtained by considering only local node properties, such as degree distributions. Many networks, however, possess large-scale modular structures that can drastically influence their spectral characteristics and which are neglected in such simplified descriptions. Here, we obtain in a unified fashion the spectrum of a large family of operators, including the adjacency, Laplacian, and normalized Laplacian matrices, for networks with generic modular structure, in the limit of large degrees. We focus on the conditions necessary for the merging of the isolated eigenvalues with the continuous band of the spectrum, after which the planted modular structure can no longer be easily detected by spectral methods. This is a crucial transition point which determines when a modular structure is strong enough to affect a given dynamical process. We show that this transition happens in general at different points for the different matrices, and hence the detectability threshold can vary significantly, depending on the operator chosen. Equivalently, the sensitivity to the modular structure of the different dynamical processes associated with each matrix will be different, given the same large-scale structure present in the network. Furthermore, we show that, with the exception of the Laplacian matrix, the different transitions coalesce into the same point for the special case where the modules are homogeneous but separate otherwise.

  4. Identification of Polyketide Inhibitors Targeting 3-Dehydroquinate Dehydratase in the Shikimate Pathway of Enterococcus faecalis

    PubMed Central

    Hernandez-Valladares, Maria; Go, Maybelle Kho; Tung, Alvin; Aguda, Adeleke H.; Robinson, Robert C.; Yew, Wen Shan

    2014-01-01

    Due to the emergence of resistance toward current antibiotics, there is a pressing need to develop the next generation of antibiotics as therapeutics against infectious and opportunistic diseases of microbial origins. The shikimate pathway is exclusive to microbes, plants and fungi, and hence is an attractive and logical target for development of antimicrobial therapeutics. The Gram-positive commensal microbe, Enterococcus faecalis, is a major human pathogen associated with nosocomial infections and resistance to vancomycin, the “drug of last resort”. Here, we report the identification of several polyketide-based inhibitors against the E. faecalis shikimate pathway enzyme, 3-dehydroquinate dehydratase (DHQase). In particular, marein, a flavonoid polyketide, both inhibited DHQase and retarded the growth of Enterococcus faecalis. The purification, crystallization and structural resolution of recombinant DHQase from E. faecalis (at 2.2 Å resolution) are also reported. This study provides a route in the development of polyketide-based antimicrobial inhibitors targeting the shikimate pathway of the human pathogen E. faecalis. PMID:25072253

  5. Enhancement of lovastatin production by supplementing polyketide antibiotics to the submerged culture of Aspergillus terreus.

    PubMed

    Jia, Zhihua; Zhang, Xiaoli; Zhao, Yaling; Cao, Xuejun

    2010-04-01

    Feedback inhibition existed in lovastatin biosynthesis from Aspergillus terreus. Exogenous lovastatin and other different polyketide antibiotics biosynthesized by polyketide synthase were supplemented to the cultures of A. terreus to investigate their influences on lovastatin production. Supplementing exogenous lovastatin of 100 mg l(-1) at the early stage of fermentation and the fast stage of its biosynthesis resulted in decreases of 76.4% and 20% in final lovastatin production, respectively. However, the fungal cell growth was not affected; the growing cycle was only prolonged in the submerged cultivation. Separate supplementation of the five kinds of polyketide antibiotics such as tylosin, erythromycin, tetracycline, daunorobin, and rifamycin to the cultures resulted in increases of about 20 approximately 25% in the final lovastatin production. Especially, supplementing tylosin of 50 mg l(-1) at the beginning of lovastatin biosynthesis led to the final lovastatin production of 952.7 +/- 24.3 mg l(-1), which was improved by 42% and 22% compared with that produced in the control and the original culture, respectively. These results are helpful to understand the regulations on lovastatin biosynthesis and improve the final desired metabolite contents in many antibiotics production.

  6. Recognition of Acyl Carrier Proteins by Ketoreductases in Assembly Line Polyketide Synthases

    PubMed Central

    Ostrowski, Matthew P.; Cane, David E.; Khosla, Chaitan

    2016-01-01

    Ketoreductases (KRs) are the most widespread tailoring domains found in individual modules of assembly line polyketide synthases (PKSs), and are responsible for controlling the configurations of both the α-methyl and β-hydroxyl stereogenic centers in the growing polyketide chain. Because they recognize substrates that are covalently bound to acyl carrier proteins (ACPs) within the same PKS module, we sought to quantify the extent to which protein-protein recognition contributes to the turnover of these oxidoreductive enzymes using stand-alone domains from the 6-deoxyerythronolide B synthase (DEBS). Reduced 2-methyl-3-hydroxyacyl-ACP substrates derived from two enantiomeric acyl chains and four distinct ACP domains were synthesized and presented to four distinct KR domains. Two KRs, from DEBS modules 2 and 5, displayed little preference for oxidation of substrates tethered to their cognate ACP domains over those attached to the other ACP domains tested. In contrast, the KR from DEBS module 1 showed a ca. 10-50-fold preference for substrate attached to its native ACP domain, whereas the KR from DEBS module 6 actually displayed a ca. 10-fold preference for the ACP from DEBS module 5. Our findings suggest that recognition of the ACP by a KR domain is unlikely to affect the rate of native assembly line polyketide biosynthesis. In some cases, however, unfavorable KR-ACP interactions may suppress the rate of substrate processing when KR domains are swapped to construct hybrid PKS modules. PMID:27118242

  7. Unprecedented diversity of catalytic domains in the first four modules of the putative pederin polyketide synthase.

    PubMed

    Piel, Jörn; Wen, Gaiping; Platzer, Matthias; Hui, Dequan

    2004-01-03

    Polyketides of the pederin group are highly potent antitumor compounds found in terrestrial beetles and marine sponges. Pederin is used by beetles of the genera Paederus and Paederidus as a chemical defense. We have recently identified a group of putative pederin biosynthesis genes and localized them to the genome of an as yet unculturable Pseudomonas sp. symbiont, the likely true pederin producer. However, this polyketide synthase cluster lacks several genes expected for pederin production. Here we report an additional polyketide synthase encoded on a separate region of the symbiont genome. It contains at least three novel catalytic domains that are predicted to be involved in pederin chain initiation and the formation of an unusual exomethylene bond. The region is bordered by mobility pseudogenes; this suggests that gene transposition led to the disjointed cluster organization. With this work, all putative pederin genes have been identified. Their heterologous expression in a culturable bacterium will provide important insights into how sustainable sources of invertebrate-derived drug candidates can be created.

  8. Antitumor polyketide biosynthesis by an uncultivated bacterial symbiont of the marine sponge Theonella swinhoei.

    PubMed

    Piel, Jörn; Hui, Dequan; Wen, Gaiping; Butzke, Daniel; Platzer, Matthias; Fusetani, Nobuhiro; Matsunaga, Shigeki

    2004-11-16

    Bacterial symbionts have long been suspected to be the true producers of many drug candidates isolated from marine invertebrates. Sponges, the most important marine source of biologically active natural products, have been frequently hypothesized to contain compounds of bacterial origin. This symbiont hypothesis, however, remained unproven because of a general inability to cultivate the suspected producers. However, we have recently identified an uncultured Pseudomonas sp. symbiont as the most likely producer of the defensive antitumor polyketide pederin in Paederus fuscipes beetles by cloning the putative biosynthesis genes. Here we report closely related genes isolated from the highly complex metagenome of the marine sponge Theonella swinhoei, which is the source of the onnamides and theopederins, a group of polyketides that structurally resemble pederin. Sequence features of the isolated genes clearly indicate that it belongs to a prokaryotic genome and should be responsible for the biosynthesis of almost the entire portion of the polyketide structure that is correlated with antitumor activity. Besides providing further proof for the role of the related beetle symbiont-derived genes, these findings raise intriguing ecological and evolutionary questions and have important general implications for the sustainable production of otherwise inaccessible marine drugs by using biotechnological strategies.

  9. Computational identification and analysis of orphan assembly-line polyketide synthases

    PubMed Central

    O’Brien, Robert V; Davis, Ronald W; Khosla, Chaitan; Hillenmeyer, Maureen E

    2014-01-01

    The increasing availability of DNA sequence data offers an opportunity for identifying new assembly-line polyketide synthases (PKSs) that produce biologically active natural products. We developed an automated method to extract and consolidate all multimodular PKS sequences (including hybrid PKS/non-ribosomal peptide synthetases) in the National Center for Biotechnology Information (NCBI) database, generating a non-redundant catalog of 885 distinct assembly-line PKSs, the majority of which were orphans associated with no known polyketide product. Two in silico experiments highlight the value of this search method and resulting catalog. First, we identified an orphan that could be engineered to produce an analog of albocycline, an interesting antibiotic whose gene cluster has not yet been sequenced. Second, we identified and analyzed a hitherto overlooked family of metazoan multimodular PKSs, including one from Caenorhabditis elegans. We also developed a comparative analysis method that identified sequence relationships among known and orphan PKSs. As expected, PKS sequences clustered according to structural similarities between their polyketide products. The utility of this method was illustrated by highlighting an interesting orphan from the genus Burkholderia that has no close relatives. Our search method and catalog provide a community resource for the discovery of new families of assembly-line PKSs and their antibiotic products. PMID:24301183

  10. Structure of the polyketide cyclase SnoaL reveals a novel mechanism for enzymatic aldol condensation.

    PubMed

    Sultana, Azmiri; Kallio, Pauli; Jansson, Anna; Wang, Ji-Shu; Niemi, Jarmo; Mäntsälä, Pekka; Schneider, Gunter

    2004-05-05

    SnoaL belongs to a family of small polyketide cyclases, which catalyse ring closure steps in the biosynthesis of polyketide antibiotics produced in Streptomyces. Several of these antibiotics are among the most used anti-cancer drugs currently in use. The crystal structure of SnoaL, involved in nogalamycin biosynthesis, with a bound product, has been determined to 1.35 A resolution. The fold of the subunit can be described as a distorted alpha+beta barrel, and the ligand is bound in the hydrophobic interior of the barrel. The 3D structure and site-directed mutagenesis experiments reveal that the mechanism of the intramolecular aldol condensation catalysed by SnoaL is different from that of the classical aldolases, which employ covalent Schiff base formation or a metal ion cofactor. The invariant residue Asp121 acts as an acid/base catalyst during the reaction. Stabilisation of the enol(ate) intermediate is mainly achieved by the delocalisation of the electron pair over the extended pi system of the substrate. These polyketide cyclases thus form of family of enzymes with a unique catalytic strategy for aldol condensation.

  11. Structure of the polyketide cyclase SnoaL reveals a novel mechanism for enzymatic aldol condensation

    PubMed Central

    Sultana, Azmiri; Kallio, Pauli; Jansson, Anna; Wang, Ji-Shu; Niemi, Jarmo; Mäntsälä, Pekka; Schneider, Gunter

    2004-01-01

    SnoaL belongs to a family of small polyketide cyclases, which catalyse ring closure steps in the biosynthesis of polyketide antibiotics produced in Streptomyces. Several of these antibiotics are among the most used anti-cancer drugs currently in use. The crystal structure of SnoaL, involved in nogalamycin biosynthesis, with a bound product, has been determined to 1.35 Å resolution. The fold of the subunit can be described as a distorted α+β barrel, and the ligand is bound in the hydrophobic interior of the barrel. The 3D structure and site-directed mutagenesis experiments reveal that the mechanism of the intramolecular aldol condensation catalysed by SnoaL is different from that of the classical aldolases, which employ covalent Schiff base formation or a metal ion cofactor. The invariant residue Asp121 acts as an acid/base catalyst during the reaction. Stabilisation of the enol(ate) intermediate is mainly achieved by the delocalisation of the electron pair over the extended π system of the substrate. These polyketide cyclases thus form of family of enzymes with a unique catalytic strategy for aldol condensation. PMID:15071504

  12. Analysis of the Ketosynthase-Chain Length Factor Heterodimer from the Fredericamycin Polyketide Synthase

    PubMed Central

    Szu, Ping-Hui; Govindarajan, Sridhar; Meehan, Michael J.; Das, Abhirup; Nguyen, Don D.; Dorrestein, Pieter C.; Minshull, Jeremy; Khosla, Chaitan

    2011-01-01

    SUMMARY The pentadecaketide fredericamycin has the longest carbon chain backbone among polycyclic aromatic polyketide antibiotics whose biosynthetic genes have been sequenced. This backbone is synthesized by the bimodular fdm polyketide synthase (PKS). The initiation module is thought to synthesize a C6 intermediate that is then transferred onto the elongation PKS module, which extends it into a C30 poly-β-ketoacyl product. Here we demonstrate that the bimodular fdm PKS as well as its elongation module alone synthesize undecaketides and dodecaketides. Thus, unlike other homologues, the fdm ketosynthase – chain length factor (KS-CLF) heterodimer does not exclusively control the backbone length of its natural product. Using sequence- and structure-based approaches, 48 multiple mutants of the CLF were engineered and analyzed. Unexpectedly, the I134F mutant was unable to turn over, but could initiate and at least partially elongate the polyketide chain. This unprecedented mutant suggests that the KS-CLF heterodimer harbors an as yet uncharacterized chain termination mechanism. Together, our findings reveal fundamental mechanistic differences between the fdm PKS and its well-studied homologues. PMID:21867917

  13. Pseudomonas syringae Phytotoxins: Mode of Action, Regulation, and Biosynthesis by Peptide and Polyketide Synthetases

    PubMed Central

    Bender, Carol L.; Alarcón-Chaidez, Francisco; Gross, Dennis C.

    1999-01-01

    Coronatine, syringomycin, syringopeptin, tabtoxin, and phaseolotoxin are the most intensively studied phytotoxins of Pseudomonas syringae, and each contributes significantly to bacterial virulence in plants. Coronatine functions partly as a mimic of methyl jasmonate, a hormone synthesized by plants undergoing biological stress. Syringomycin and syringopeptin form pores in plasma membranes, a process that leads to electrolyte leakage. Tabtoxin and phaseolotoxin are strongly antimicrobial and function by inhibiting glutamine synthetase and ornithine carbamoyltransferase, respectively. Genetic analysis has revealed the mechanisms responsible for toxin biosynthesis. Coronatine biosynthesis requires the cooperation of polyketide and peptide synthetases for the assembly of the coronafacic and coronamic acid moieties, respectively. Tabtoxin is derived from the lysine biosynthetic pathway, whereas syringomycin, syringopeptin, and phaseolotoxin biosynthesis requires peptide synthetases. Activation of phytotoxin synthesis is controlled by diverse environmental factors including plant signal molecules and temperature. Genes involved in the regulation of phytotoxin synthesis have been located within the coronatine and syringomycin gene clusters; however, additional regulatory genes are required for the synthesis of these and other phytotoxins. Global regulatory genes such as gacS modulate phytotoxin production in certain pathovars, indicating the complexity of the regulatory circuits controlling phytotoxin synthesis. The coronatine and syringomycin gene clusters have been intensively characterized and show potential for constructing modified polyketides and peptides. Genetic reprogramming of peptide and polyketide synthetases has been successful, and portions of the coronatine and syringomycin gene clusters could be valuable resources in developing new antimicrobial agents. PMID:10357851

  14. Predicted incorporation of non-native substrates by a polyketide synthase yields bioactive natural product derivatives.

    PubMed

    Bravo-Rodriguez, Kenny; Ismail-Ali, Ahmed F; Klopries, Stephan; Kushnir, Susanna; Ismail, Shehab; Fansa, Eyad K; Wittinghofer, Alfred; Schulz, Frank; Sanchez-Garcia, Elsa

    2014-09-05

    The polyether ionophore monensin is biosynthesized by a polyketide synthase that delivers a mixture of monensins A and B by the incorporation of ethyl- or methyl-malonyl-CoA at its fifth module. Here we present the first computational model of the fifth acyltransferase domain (AT5mon ) of this polyketide synthase, thus affording an investigation of the basis of the relaxed specificity in AT5mon , insights into the activation for the nucleophilic attack on the substrate, and prediction of the incorporation of synthetic malonic acid building blocks by this enzyme. Our predictions are supported by experimental studies, including the isolation of a predicted derivative of the monensin precursor premonensin. The incorporation of non-native building blocks was found to alter the ratio of premonensins A and B. The bioactivity of the natural product derivatives was investigated and revealed binding to prenyl-binding protein. We thus show the potential of engineered biosynthetic polyketides as a source of ligands for biological macromolecules. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  15. A new reducing polyketide synthase gene from the lichen-forming fungus Cladonia metacorallifera.

    PubMed

    Kim, Jung A; Hong, Soon Gyu; Cheong, Yong Hwa; Koh, Young Jin; Hur, Jae-Seoun

    2012-01-01

    Lichens produce unique polyketide secondary metabolites including depsides, depsidones, dibenzofurans and depsones. The biosynthesis of these compounds is governed by polyketide synthase (PKS), but the mechanism via which they are produced has remained unclear until now. We reported the 6-methylsalicylic acid synthase (6-MSAS) type of PKS gene, which is a member of the fungal-reducing PKSs. A cultured mycobiont of Cladonia metacorallifera was employed in the isolation and characterization of a polyketide synthase gene (CmPKS1). The complete sequence information for CmPKS1 was acquired via the screening of a Fosmid genomic library with a 456 bp fragment corresponding to part of the acyl transferase (AT) domain as a probe. CmPKS1 contains β-ketoacyl synthase (KS), AT, dehydratase (DH), ketoreductase (KR) and phosphopantetheine attachment site (PP) domains.: The domain organization of CmPKS1 (KS-AT-DH-KR-PP) is a typical 6-MSAS-type PKS, and the results of phylogenetic analysis showed that CmPKS1 grouped with other fungal-reducing PKSs. Quantitative real time PCR analyses showed that CmPKS1 was expressed preferentially in the early growth stage of the axenically cultured mycobiont. Furthermore CmPKS1 expression was found to be dependent on the carbon sources and concentrations in the medium.

  16. Computational identification and analysis of orphan assembly-line polyketide synthases.

    PubMed

    O'Brien, Robert V; Davis, Ronald W; Khosla, Chaitan; Hillenmeyer, Maureen E

    2014-01-01

    The increasing availability of DNA sequence data offers an opportunity for identifying new assembly-line polyketide synthases (PKSs) that produce biologically active natural products. We developed an automated method to extract and consolidate all multimodular PKS sequences (including hybrid PKS/non-ribosomal peptide synthetases) in the National Center for Biotechnology Information (NCBI) database, generating a non-redundant catalog of 885 distinct assembly-line PKSs, the majority of which were orphans associated with no known polyketide product. Two in silico experiments highlight the value of this search method and resulting catalog. First, we identified an orphan that could be engineered to produce an analog of albocycline, an interesting antibiotic whose gene cluster has not yet been sequenced. Second, we identified and analyzed a hitherto overlooked family of metazoan multimodular PKSs, including one from Caenorhabditis elegans. We also developed a comparative analysis method that identified sequence relationships among known and orphan PKSs. As expected, PKS sequences clustered according to structural similarities between their polyketide products. The utility of this method was illustrated by highlighting an interesting orphan from the genus Burkholderia that has no close relatives. Our search method and catalog provide a community resource for the discovery of new families of assembly-line PKSs and their antibiotic products.

  17. Product modular design incorporating preventive maintenance issues

    NASA Astrophysics Data System (ADS)

    Gao, Yicong; Feng, Yixiong; Tan, Jianrong

    2016-03-01

    Traditional modular design methods lead to product maintenance problems, because the module form of a system is created according to either the function requirements or the manufacturing considerations. For solving these problems, a new modular design method is proposed with the considerations of not only the traditional function related attributes, but also the maintenance related ones. First, modularity parameters and modularity scenarios for product modularity are defined. Then the reliability and economic assessment models of product modularity strategies are formulated with the introduction of the effective working age of modules. A mathematical model used to evaluate the difference among the modules of the product so that the optimal module of the product can be established. After that, a multi-objective optimization problem based on metrics for preventive maintenance interval different degrees and preventive maintenance economics is formulated for modular optimization. Multi-objective GA is utilized to rapidly approximate the Pareto set of optimal modularity strategy trade-offs between preventive maintenance cost and preventive maintenance interval difference degree. Finally, a coordinate CNC boring machine is adopted to depict the process of product modularity. In addition, two factorial design experiments based on the modularity parameters are constructed and analyzed. These experiments investigate the impacts of these parameters on the optimal modularity strategies and the structure of module. The research proposes a new modular design method, which may help to improve the maintainability of product in modular design.

  18. Modular process modeling for OPC

    NASA Astrophysics Data System (ADS)

    Keck, M. C.; Bodendorf, C.; Schmidtling, T.; Schlief, R.; Wildfeuer, R.; Zumpe, S.; Niehoff, M.

    2007-03-01

    Modular OPC modeling, describing mask, optics, resist and etch processes separately is an approach to keep efforts for OPC manageable. By exchanging single modules of a modular OPC model, a fast response to process changes during process development is possible. At the same time efforts can be reduced, since only single modular process steps have to be re-characterized as input for OPC modeling as the process is adjusted and optimized. Commercially available OPC tools for full chip processing typically make use of semi-empirical models. The goal of our work is to investigate to what extent these OPC tools can be applied for modeling of single process steps as separate modules. For an advanced gate level process we analyze the modeling accuracy over different process conditions (focus and dose) when combining models for each process step - optics, resist and etch - for differing single processes to a model describing the total process.

  19. Modular assembly of optical nanocircuits.

    PubMed

    Shi, Jinwei; Monticone, Francesco; Elias, Sarah; Wu, Yanwen; Ratchford, Daniel; Li, Xiaoqin; Alù, Andrea

    2014-05-29

    A key element enabling the microelectronic technology advances of the past decades has been the conceptualization of complex circuits with versatile functionalities as being composed of the proper combination of basic 'lumped' circuit elements (for example, inductors and capacitors). In contrast, modern nanophotonic systems are still far from a similar level of sophistication, partially because of the lack of modularization of their response in terms of basic building blocks. Here we demonstrate the design, assembly and characterization of relatively complex photonic nanocircuits by accurately positioning a number of metallic and dielectric nanoparticles acting as modular lumped elements. The nanoparticle clusters produce the desired spectral response described by simple circuit rules and are shown to be dynamically reconfigurable by modifying the direction or polarization of impinging signals. Our work represents an important step towards extending the powerful modular design tools of electronic circuits into nanophotonic systems.

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

    PubMed

    Yang, Xinmei; Matsui, Takashi; Kodama, Takeshi; Mori, Takahiro; Zhou, Xiaoxi; Taura, Futoshi; Noguchi, Hiroshi; Abe, Ikuro; Morita, Hiroyuki

    2016-03-01

    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 α+β barrel (DABB) protein that catalyzes the C2-C7 aldol cyclization of the linear pentyl tetra-β-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 1.70 Å 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. Structural data reported in this paper are available in the Protein Data Bank under the accession numbers 5B08 for the OAC apo, 5B09 for the OAC-OA binary complex and 5B0A, 5B0B, 5B0C, 5B0D, 5B0E, 5B0F and 5B0G for the OAC His5Q, Ile7F, Tyr27F, Tyr27W, Val59M, Tyr72F and His78S mutant enzymes, respectively. © 2016 Federation of European Biochemical Societies.

  1. Modular multichannel surface plasmon spectrometer

    NASA Astrophysics Data System (ADS)

    Neuert, G.; Kufer, S.; Benoit, M.; Gaub, H. E.

    2005-05-01

    We have developed a modular multichannel surface plasmon resonance (SPR) spectrometer on the basis of a commercially available hybrid sensor chip. Due to its modularity this inexpensive and easy to use setup can readily be adapted to different experimental environments. High temperature stability is achieved through efficient thermal coupling of individual SPR units. With standard systems the performance of the multichannel instrument was evaluated. The absorption kinetics of a cysteamine monolayer, as well as the concentration dependence of the specific receptor-ligand interaction between biotin and streptavidin was measured.

  2. Modular multivariable control improves hydrocracking

    SciTech Connect

    Chia, T.L.; Lefkowitz, I.; Tamas, P.D.

    1996-10-01

    Modular multivariable control (MMC), a system of interconnected, single process variable controllers, can be a user-friendly, reliable and cost-effective alternative to centralized, large-scale multivariable control packages. MMC properties and features derive directly from the properties of the coordinated controller which, in turn, is based on internal model control technology. MMC was applied to a hydrocracking unit involving two process variables and three controller outputs. The paper describes modular multivariable control, MMC properties, tuning considerations, application at the DCS level, constraints handling, and process application and results.

  3. Inherent controllability in modular ALMRs

    SciTech Connect

    Sackett, J.I.; Sevy, R.H.; Wei, T.Y.C.

    1989-01-01

    As part of recent development efforts on advanced reactor designs ANL has proposed the IFR (Integral Fast Reactor) concept. The IFR concept is currently being applied to modular sized reactors which would be built in multiple power paks together with an integrated fuel cycle facility. It has been amply demonstrated that the concept as applied to the modular designs has significant advantages in regard to ATWS transients. Attention is now being focussed on determining whether or not those advantages deriving from the traits of the IFR can be translated to the operational/DBA (design basis accident) class of transients. 5 refs., 3 figs., 3 tabs.

  4. Modular Firewalls for Storage Areas

    NASA Technical Reports Server (NTRS)

    Fedor, O. H.; Owens, L. J.

    1986-01-01

    Giant honeycomb structures assembled in modular units. Flammable materials stored in cells. Walls insulated with firebrick to prevent spread of fire among cells. Portable, modular barrier withstands heat of combustion for limited time and confines combustion products horizontally to prevent fire from spreading. Barrier absorbs heat energy by ablation and not meant to be reused. Designed to keep fires from spreading among segments of solid rocket propellant in storage, barrier erected between storage units of other flammable or explosive materials; tanks of petroleum or liquid natural gas. Barrier adequate for most industrial purposes.

  5. Modularity in Cognition: Framing the Debate

    ERIC Educational Resources Information Center

    Barrett, H. Clark; Kurzban, Robert

    2006-01-01

    Modularity has been the subject of intense debate in the cognitive sciences for more than 2 decades. In some cases, misunderstandings have impeded conceptual progress. Here the authors identify arguments about modularity that either have been abandoned or were never held by proponents of modular views of the mind. The authors review arguments that…

  6. 47 CFR 15.212 - Modular transmitters.

    Code of Federal Regulations, 2014 CFR

    2014-10-01

    ... 47 Telecommunication 1 2014-10-01 2014-10-01 false Modular transmitters. 15.212 Section 15.212 Telecommunication FEDERAL COMMUNICATIONS COMMISSION GENERAL RADIO FREQUENCY DEVICES Intentional Radiators § 15.212 Modular transmitters. (a) Single modular transmitters consist of a completely...

  7. 47 CFR 15.212 - Modular transmitters.

    Code of Federal Regulations, 2010 CFR

    2010-10-01

    ... 47 Telecommunication 1 2010-10-01 2010-10-01 false Modular transmitters. 15.212 Section 15.212 Telecommunication FEDERAL COMMUNICATIONS COMMISSION GENERAL RADIO FREQUENCY DEVICES Intentional Radiators § 15.212 Modular transmitters. (a) Single modular transmitters consist of a completely...

  8. 47 CFR 15.212 - Modular transmitters.

    Code of Federal Regulations, 2013 CFR

    2013-10-01

    ... 47 Telecommunication 1 2013-10-01 2013-10-01 false Modular transmitters. 15.212 Section 15.212 Telecommunication FEDERAL COMMUNICATIONS COMMISSION GENERAL RADIO FREQUENCY DEVICES Intentional Radiators § 15.212 Modular transmitters. (a) Single modular transmitters consist of a completely...

  9. 47 CFR 15.212 - Modular transmitters.

    Code of Federal Regulations, 2011 CFR

    2011-10-01

    ... 47 Telecommunication 1 2011-10-01 2011-10-01 false Modular transmitters. 15.212 Section 15.212 Telecommunication FEDERAL COMMUNICATIONS COMMISSION GENERAL RADIO FREQUENCY DEVICES Intentional Radiators § 15.212 Modular transmitters. (a) Single modular transmitters consist of a completely...

  10. 48 CFR 3417.70 - Modular contracting.

    Code of Federal Regulations, 2013 CFR

    2013-10-01

    ... 48 Federal Acquisition Regulations System 7 2013-10-01 2012-10-01 true Modular contracting. 3417... REGULATION CONTRACTING METHODS AND CONTRACT TYPES SPECIAL CONTRACTING METHODS Modular Contracting 3417.70 Modular contracting. (a) FSA—May incrementally conduct successive procurements of modules of overall...

  11. The Modular Mind and Intrapersonal Communication Processes.

    ERIC Educational Resources Information Center

    Stacks, Don W.

    Based on a prior model on modularity of the brain, a new modular model of intrapersonal communication was developed which focuses on brain processing, encompassing both the structures and the functions of those structures in the creation of messages. The modular mind is a bio-social model of communication which presupposes a relationship between…

  12. Evolution and the Modularity of Mindreading.

    ERIC Educational Resources Information Center

    Moore, Chris

    1996-01-01

    Reviews Baron-Cohen's study of autism and an explanatory theory called modularity of mindreading, which proposed a domain-specific modular psychological model based on evolutionary, developmental, psychopathological, and neurobiological considerations. Enumerates problems with the modularity approach and emphasized the evolution of domain general…

  13. Modular Instruction Under Restricted Conditions.

    ERIC Educational Resources Information Center

    Utomo, Tjipto; Ruijter, Kees

    1984-01-01

    Describes the evaluation and reconstruction of a transport phenomena course given at the Bandung Institute of Technology which had a 70 percent failure rate. Discusses the teacher-paced modular instruction technique designed to replace the original course material and its results in terms of student performance over a three-year period. (JM)

  14. Rapidly Deployed Modular Telemetry System

    NASA Technical Reports Server (NTRS)

    Varnavas, Kosta A. (Inventor); Sims, William Herbert, III (Inventor)

    2013-01-01

    The present invention is a telemetry system, and more specifically is a rapidly deployed modular telemetry apparatus which utilizes of SDR technology and the FPGA programming capability to reduce the number of hardware components and programming required to deploy a telemetry system.

  15. Polyketide Derivatives from Annona muricata Linn Leaves as Potencial Anticancer Material by Combination Treatment With Doxorubicin on Hela Cell Line

    NASA Astrophysics Data System (ADS)

    Artanti, A. N.; Astirin, O. P.; Prayito, A.; Widiyaningsih, R. F.; Prihapsara, F.

    2017-02-01

    One of the compounds found effication as an anticancer agent on cervical cancer is acetogenin, a polyketide compound that is abundant in Annona muricata L. leaves. This study has been done to examine polyketide derivatives was isolated from Annona muricata L. which has potency to induce apoptosis by p53 expression on hela cell line. An approach recently develop to overcome side effect of chemoterapeutic agent is used of combined chemoterapeutic agent, i.e doxorubicin. The determination of cytotoxic combination activity from polyketide derivative and doxorubicin was evaluated using MTT assay to obtain the value of CI (combination index). The expression of p53 profile was evaluated by immunohistochemistry on hela cell line. Data analysis showed that combination of polyketide derivative from Annona muricata L. (38,5 µg/ml) and doxorubicin with all of concentration performed synergistic effect on hela cell line with CI value from 0,33 - 0,65. The analysis on immucytochemistry showed that polyketide derivative from Annona muricata L. leaves could enhance p53 pathway significantly on hela cell line.

  16. Characterization and engineering of the ethylmalonyl-CoA pathway towards the improved heterologous production of polyketides in Streptomyces venezuelae.

    PubMed

    Jung, Won Seok; Kim, Eunji; Yoo, Young Ji; Ban, Yeon Hee; Kim, Eun Ji; Yoon, Yeo Joon

    2014-04-01

    Streptomyces venezuelae has an inherent advantage as a heterologous host for polyketide production due to its fast rate of growth that cannot be endowed easily through metabolic engineering. However, the utility of S. venezuelae as a host has been limited thus far due to its inadequate intracellular reserves of the (2S)-ethylmalonyl-CoA building block needed to support the biosynthesis of polyketides preventing the efficient production of the desired metabolite, such as tylactone. Here, via precursor supply engineering, we demonstrated that S. venezuelae can be developed into a more efficient general heterologous host for the quick production of polyketides. We first identified and functionally characterized the ethylmalonyl-CoA pathway which plays a major role in supplying the (2S)-ethylmalonyl-CoA extender unit in S. venezuelae. Next, S. venezuelae was successfully engineered to increase the intracellular ethylmalonyl-CoA concentration by the deletion of the meaA gene encoding coenzyme B₁₂-dependent ethylmalonyl-CoA mutase in combination with ethylmalonate supplementation and was engineered to upregulate the expression of the heterologous tylosin PKS by overexpression of the pathway specific regulatory gene pikD. Thus, a dramatic increase (∼10-fold) in tylactone production was achieved. In addition, the detailed insights into the role of the ethylmalonyl-CoA pathway, which is present in most streptomycetes, provides a general strategy to increase the ethylmalonyl-CoA supply for polyketide biosynthesis in the most prolific family of polyketide-producing bacteria.

  17. ABCG26-Mediated Polyketide Trafficking and Hydroxycinnamoyl Spermidines Contribute to Pollen Wall Exine Formation in Arabidopsis[W

    PubMed Central

    Quilichini, Teagen D.; Samuels, A. Lacey; Douglas, Carl J.

    2014-01-01

    Pollen grains are encased by a multilayered, multifunctional wall. The sporopollenin and pollen coat constituents of the outer pollen wall (exine) are contributed by surrounding sporophytic tapetal cells. Because the biosynthesis and development of the exine occurs in the innermost cell layers of the anther, direct observations of this process are difficult. The objective of this study was to investigate the transport and assembly of exine components from tapetal cells to microspores in the intact anthers of Arabidopsis thaliana. Intrinsically fluorescent components of developing tapetum and microspores were imaged in intact, live anthers using two-photon microscopy. Mutants of ABCG26, which encodes an ATP binding cassette transporter required for exine formation, accumulated large fluorescent vacuoles in tapetal cells, with corresponding loss of fluorescence on microspores. These vacuolar inclusions were not observed in tapetal cells of double mutants of abcg26 and genes encoding the proposed sporopollenin polyketide biosynthetic metabolon (ACYL COENZYME A SYNTHETASE5, POLYKETIDE SYNTHASE A [PKSA], PKSB, and TETRAKETIDE α-PYRONE REDUCTASE1), providing a genetic link between transport by ABCG26 and polyketide biosynthesis. Genetic analysis also showed that hydroxycinnamoyl spermidines, known components of the pollen coat, were exported from tapeta prior to programmed cell death in the absence of polyketides, raising the possibility that they are incorporated into the exine prior to pollen coat deposition. We propose a model where ABCG26-exported polyketides traffic from tapetal cells to form the sporopollenin backbone, in coordination with the trafficking of additional constituents, prior to tapetum programmed cell death. PMID:25415974

  18. Quasispecies theory for evolution of modularity

    NASA Astrophysics Data System (ADS)

    Park, Jeong-Man; Niestemski, Liang Ren; Deem, Michael W.

    2015-01-01

    Biological systems are modular, and this modularity evolves over time and in different environments. A number of observations have been made of increased modularity in biological systems under increased environmental pressure. We here develop a quasispecies theory for the dynamics of modularity in populations of these systems. We show how the steady-state fitness in a randomly changing environment can be computed. We derive a fluctuation dissipation relation for the rate of change of modularity and use it to derive a relationship between rate of environmental changes and rate of growth of modularity. We also find a principle of least action for the evolved modularity at steady state. Finally, we compare our predictions to simulations of protein evolution and find them to be consistent.

  19. Quasispecies Theory for Evolution of Modularity

    PubMed Central

    Park, Jeong-Man; Niestemski, Liang Ren; Deem, Michael W.

    2015-01-01

    Biological systems are modular, and this modularity evolves over time and in different environments. A number of observations have been made of increased modularity in biological systems under increased environmental pressure. We here develop a quasispecies theory for the dynamics of modularity in populations of these systems. We show how the steady-state fitness in a randomly changing environment can be computed. We derive a fluctuation dissipation relation for the rate of change of modularity and use it to derive a relationship between rate of environmental changes and rate of growth of modularity. We also find a principle of least action for the evolved modularity at steady state. Finally, we compare our predictions to simulations of protein evolution and find them to be consistent. PMID:25679649

  20. Quasispecies theory for evolution of modularity.

    PubMed

    Park, Jeong-Man; Niestemski, Liang Ren; Deem, Michael W

    2015-01-01

    Biological systems are modular, and this modularity evolves over time and in different environments. A number of observations have been made of increased modularity in biological systems under increased environmental pressure. We here develop a quasispecies theory for the dynamics of modularity in populations of these systems. We show how the steady-state fitness in a randomly changing environment can be computed. We derive a fluctuation dissipation relation for the rate of change of modularity and use it to derive a relationship between rate of environmental changes and rate of growth of modularity. We also find a principle of least action for the evolved modularity at steady state. Finally, we compare our predictions to simulations of protein evolution and find them to be consistent.

  1. Studies of the production of fungal polyketides in Aspergillus nidulans by using systems biology tools.

    PubMed

    Panagiotou, Gianni; Andersen, Mikael R; Grotkjaer, Thomas; Regueira, Torsten B; Nielsen, Jens; Olsson, Lisbeth

    2009-04-01

    Many filamentous fungi produce polyketide molecules with great significance as human pharmaceuticals; these molecules include the cholesterol-lowering compound lovastatin, which was originally isolated from Aspergillus terreus. The chemical diversity and potential uses of these compounds are virtually unlimited, and it is thus of great interest to develop a well-described microbial production platform for polyketides. Using genetic engineering tools available for the model organism Aspergillus nidulans, we constructed two recombinant strains, one expressing the Penicillium griseofulvum 6-methylsalicylic acid (6-MSA) synthase gene and one expressing the 6-MSA synthase gene and overexpressing the native xylulose-5-phosphate phosphoketolase gene (xpkA) for increasing the pool of polyketide precursor levels. The physiology of the recombinant strains and that of a reference wild-type strain were characterized on glucose, xylose, glycerol, and ethanol media in controlled bioreactors. Glucose was found to be the preferred carbon source for 6-MSA production, and 6-MSA concentrations up to 455 mg/liter were obtained for the recombinant strain harboring the 6-MSA gene. Our findings indicate that overexpression of xpkA does not directly improve 6-MSA production on glucose, but it is possible, if the metabolic flux through the lower part of glycolysis is reduced, to obtain quite high yields for conversion of sugar to 6-MSA. Systems biology tools were employed for in-depth analysis of the metabolic processes. Transcriptome analysis of 6-MSA-producing strains grown on glucose and xylose in the presence and absence of xpkA overexpression, combined with flux and physiology data, enabled us to propose an xpkA-msaS interaction model describing the competition between biomass formation and 6-MSA production for the available acetyl coenzyme A.

  2. RNA Sequencing Revealed Numerous Polyketide Synthase Genes in the Harmful Dinoflagellate Karenia mikimotoi

    PubMed Central

    Kimura, Kei; Okuda, Shujiro; Nakayama, Kei; Shikata, Tomoyuki; Takahashi, Fumio; Yamaguchi, Haruo; Skamoto, Setsuko; Yamaguchi, Mineo; Tomaru, Yuji

    2015-01-01

    The dinoflagellate Karenia mikimotoi forms blooms in the coastal waters of temperate regions and occasionally causes massive fish and invertebrate mortality. This study aimed to elucidate the toxic effect of K. mikimotoi on marine organisms by using the genomics approach; RNA-sequence libraries were constructed, and data were analyzed to identify toxin-related genes. Next-generation sequencing produced 153,406 transcript contigs from the axenic culture of K. mikimotoi. BLASTX analysis against all assembled contigs revealed that 208 contigs were polyketide synthase (PKS) sequences. Thus, K. mikimotoi was thought to have several genes encoding PKS metabolites and to likely produce toxin-like polyketide molecules. Of all the sequences, approximately 30 encoded eight PKS genes, which were remarkably similar to those of Karenia brevis. Our phylogenetic analyses showed that these genes belonged to a new group of PKS type-I genes. Phylogenetic and active domain analyses showed that the amino acid sequence of four among eight Karenia PKS genes was not similar to any of the reported PKS genes. These PKS genes might possibly be associated with the synthesis of polyketide toxins produced by Karenia species. Further, a homology search revealed 10 contigs that were similar to a toxin gene responsible for the synthesis of saxitoxin (sxtA) in the toxic dinoflagellate Alexandrium fundyense. These contigs encoded A1–A3 domains of sxtA genes. Thus, this study identified some transcripts in K. mikimotoi that might be associated with several putative toxin-related genes. The findings of this study might help understand the mechanism of toxicity of K. mikimotoi and other dinoflagellates. PMID:26561394

  3. Effective Antibiofilm Polyketides against Staphylococcus aureus from the Pyranonaphthoquinone Biosynthetic Pathways of Streptomyces Species.

    PubMed

    Oja, Terhi; San Martin Galindo, Paola; Taguchi, Takaaki; Manner, Suvi; Vuorela, Pia M; Ichinose, Koji; Metsä-Ketelä, Mikko; Fallarero, Adyary

    2015-10-01

    Streptomyces bacteria are renowned for their ability to produce bioactive secondary metabolites. Recently, synthetic biology has enabled the production of intermediates and shunt products, which may have altered biological activities compared to the end products of the pathways. Here, we have evaluated the potential of recently isolated alnumycins and other closely related pyranonaphthoquinone (PNQ) polyketides against Staphylococcus aureus biofilms. The antimicrobial potency of the compounds against planktonic cells and biofilms was determined by redox dye-based viability staining, and the antibiofilm efficacy of the compounds was confirmed by viable counting. A novel antistaphylococcal polyketide, alnumycin D, was identified. Unexpectedly, the C-ribosylated pathway shunt product alnumycin D was more active against planktonic and biofilm cells than the pathway end product alnumycin A, where a ribose unit has been converted into a dioxane moiety. The evaluation of the antibiofilm potential of other alnumycins revealed that the presence of the ribose moiety in pyranose form is essential for high activity against preformed biofilms. Furthermore, the antibiofilm potential of other closely related PNQ polyketides was examined. Based on their previously reported activity against planktonic S. aureus cells, granaticin B, kalafungin, and medermycin were also selected for testing, and among them, granaticin B was found to be the most potent against preformed biofilms. The most active antibiofilm PNQs, alnumycin D and granaticin B, share several structural features that may be important for their antibiofilm activity. They are uncharged, glycosylated, and also contain a similar oxygenation pattern of the lateral naphthoquinone ring. These findings highlight the potential of antibiotic biosynthetic pathways as a source of effective antibiofilm compounds.

  4. Forazoline A: Marine-derived polyketide with antifungal in vivo efficacy

    DOE PAGES

    Wyche, Thomas P.; Piotrowski, Jeff S.; Hou, Yanpeng; ...

    2014-09-04

    Forazoline A, a novel antifungal polyketide with in vivo efficacy against Candida albicans, was discovered using LCMS-based metabolomics to investigate marine-invertebrate-associated bacteria. Forazoline A had a highly unusual and unprecedented skeleton. Acquisition of 13C–13C gCOSY and 13C–15N HMQC NMR data provided the direct carbon–carbon and carbon–nitrogen connectivity, respectively. This approach represents the first example of determining direct 13C–15N connectivity for a natural product. In this paper, using yeast chemical genomics, we propose that forazoline A operated through a new mechanism of action with a phenotypic outcome of disrupting membrane integrity.

  5. Polyketides with Immunosuppressive Activities from Mangrove Endophytic Fungus Penicillium sp. ZJ-SY2

    PubMed Central

    Liu, Hongju; Chen, Senhua; Liu, Weiyang; Liu, Yayue; Huang, Xishan; She, Zhigang

    2016-01-01

    Nine polyketides, including two new benzophenone derivatives, peniphenone (1) and methyl peniphenone (2), along with seven known xanthones (3–9) were obtained from mangrove endophytic fungus Penicillium sp. ZJ-SY2 isolated from the leaves of Sonneratia apetala. Their structures were elucidated on the basis of MS, 1D, and 2D NMR data. Compounds 1, 3, 5, and 7 showed potent immunosuppressive activity with IC50 values ranging from 5.9 to 9.3 μg/mL. PMID:27897975

  6. Coibacins A-D, Anti-leishmanial Marine Cyanobacterial Polyketides with Intriguing Biosynthetic Origins

    PubMed Central

    Balunas, Marcy J.; Grosso, Manuel F.; Villa, Francisco A.; Engene, Niclas; McPhail, Kerry L.; Tidgewell, Kevin; Pineda, Laura M.; Gerwick, Lena; Spadafora, Carmenza; Kyle, Dennis E.; Gerwick, William H.

    2012-01-01

    Four unsaturated polyketide lactone derivatives, coibacins A-D, were isolated from a Panamanian marine cyanobacterium, cf. Oscillatoria sp. The two different types of termini observed in these co-occurring metabolites, either a methyl cyclopropyl ring as seen in curacin A or a methyl vinyl chloride similar to that observed in the jamaicamides, suggest an intriguing flexibility in the “beta branch” forming biosynthetic process. The coibacins possess selective anti-leishmanial activity as well as potent anti-inflammatory activity. PMID:22794317

  7. Macrolide biosynthesis. 7. Incorporation of polyketide chain elongation intermediates into methymycin

    SciTech Connect

    Cane, D.E.; Lambalot, R.H.; Prabhakaran, P.C.; Ott, W.R. )

    1993-01-27

    Administration of [1-[sup 13]C]propionate to cultures of Streptomyces venezuelae SC 2366 gave methymycin (1), which was shown by [sup 13]C NMR analysis to be labeled at the predicted sites, C-1, C-3, C-5, C-9, and C-11. Similarly, incorporation of [1,2-[sup 13]C]acetate gave methymycin labeled at C-7 and C-8. A series of presumptive intermediates of polyketide chain elongation was also successfully incorporated. Thus, feeding of (2S,3R)-[2,3-[sup 13]C[sub 2

  8. The marine polyketide myriaporone 3/4 stalls translation by targeting the elongation phase.

    PubMed

    Muthukumar, Yazh; Roy, Myriam; Raja, Aruna; Taylor, Richard E; Sasse, Florenz

    2013-01-21

    Myriaporone 3/4, a cytotoxic polyketide, has been reported as an inhibitor of eukaryotic protein synthesis. However, the mechanism by which it inhibits translation was unknown. Here we show that myriaporone 3/4 stalls protein synthesis in the elongation phase by inducing phosphorylation of eukaryotic elongation factor 2. The phosphorylation results from direct binding of myriaporone 3/4 to eukaryotic elongation factor 2 kinase. Our study also shows that myriaporone 3/4 in the nanomolar range inhibits in vitro tube formation by endothelial cells without being cytotoxic. In general, myriaporone 3/4 was at least 300 times less toxic to primary cells than to tumor cells.

  9. Biosynthetic origin of butyrolactol A, an antifungal polyketide produced by a marine-derived Streptomyces

    PubMed Central

    Harunari, Enjuro; Komaki, Hisayuki

    2017-01-01

    Butyrolactol A is an antifungal polyketide of Streptomyces bearing an uncommon tert-butyl starter unit and a polyol system in which eight hydroxy/acyloxy carbons are contiguously connected. Except for its congener butyrolactol B, there exist no structurally related natural products to date. In this study, inspired by our previous genomic analysis, incorporation of 13C- and 2H-labeled precursors into butyrolactol A was investigated. Based on the labeling pattern and sequencing analytical data, we confirmed that the tert-butyl group is derived from valine and its C-methylation with methionine and the polyol carbons are derived from a glycolysis intermediate, possibly hydroxymalonyl-ACP. PMID:28382182

  10. Modular Stellarator Fusion Reactor concept

    SciTech Connect

    Miller, R.L.; Krakowski, R.A.

    1981-08-01

    A preliminary conceptual study is made of the Modular Stellarator Reactor (MSR). A steady-state ignited, DT-fueled, magnetic fusion reactor is proposed for use as a central electric-power station. The MSR concept combines the physics of the classic stellarator confinement topology with an innovative, modular-coil design. Parametric tradeoff calculations are described, leading to the selection of an interim design point for a 4-GWt plant based on Alcator transport scaling and an average beta value of 0.04 in an l = 2 system with a plasma aspect ratio of 11. The physics basis of the design point is described together with supporting magnetics, coil-force, and stress computations. The approach and results presented herein will be modified in the course of ongoing work to form a firmer basis for a detailed conceptual design of the MSR.

  11. Relaxation labeling using modular operators

    SciTech Connect

    Duncan, J.S.; Frei, W.

    1983-01-01

    Probabilistic relaxation labeling has been shown to be useful in image processing, pattern recognition, and artificial intelligence. The approaches taken to date have been encumbered with computationally extensive summations which generally prevent real-time operation and/or easy hardware implementation. The authors present a new and unique approach to the relaxation labeling problem using modular, VLSI-oriented hierarchical complex operators. One of the fundamental concepts of this work is the representation of the probability distribution of the possible labels for a given object (pixel) as an ellipse, which may be summed with neighboring object's distribution ellipses, resulting in a new, relaxed label space. The mathematical development of the elliptical approach will be presented and compared to more classical approaches, and a hardware block diagram that shows the implementation of the relaxation scheme using vlsi chips will be presented. Finally, results will be shown which illustrate applications of the modular scheme, iteratively, to both edges and lines. 13 references.

  12. Modular hydrodam: concept definition study

    SciTech Connect

    Not Available

    1981-07-01

    The purpose of this investigation was to explore the potential for developing economical new ultra low-head (6 to 10 ft) sites using an innovative concept known as the Modular Hydrodam (MH). This concept combines the benefits of shop fabrication, installation of equipment in truck transportable, waterproof power modules, and prefabricated gate sections that can be located between the power modules. The size and weight of the power module permits it to be fully assembled and checked out in the manufacturer's shop. The module can then be broken down into four pieces and shipped by truck to the site. Once in place, concrete ballast will be added, as necessary, to prevent flotation. The following aspects were investigated: tubular and cross flow turbines; modularized components; the use of a cable support system for horizontal stability of the dam and powerhouse; and construction in the wet as well as in the dry.

  13. Modular Platforms for Optofluidic Systems

    NASA Astrophysics Data System (ADS)

    Brammer, Marko; Mappes, Timo

    2014-01-01

    Optofluidics is increasingly gaining impact in a number of different fields of research, namely biology and medicine, environmental monitoring and green energy. However, the market for optofluidic products is still in the early development phase. In this manuscript, we discuss modular platforms as a potential concept to facilitate the transfer of optofluidic sensing systems to an industrial implementation. We present microfluidic and optical networks as a basis for the interconnection of optofluidic sensor modules. Finally, we show the potential for entire optofluidic networks

  14. PTERA - Modular Aircraft Flight Test

    NASA Image and Video Library

    2016-01-13

    Aerospace testing can be costly and time consuming but a new modular, subscale remotely piloted aircraft offers NASA researchers more affordable options for developing a wide range of cutting edge aviation and space technologies. The Prototype-Technology Evaluation and Research Aircraft (PTERA), developed by Area-I, Inc., of Kennesaw, Georgia, is an extremely versatile and high quality, yet inexpensive, flying laboratory bridging the gap between wind tunnels and crewed flight testing.

  15. Modular Platforms for Optofluidic Systems

    NASA Astrophysics Data System (ADS)

    Brammer, Marko; Mappes, Timo

    2013-02-01

    Optofluidics is increasingly gaining impact in a number of different fields of research, namely biology and medicine, environmental monitoring and green energy. However, the market for optofluidic products is still in the early development phase. In this manuscript, we discuss modular platforms as a potential concept to facilitate the transfer of optofluidic sensing systems to an industrial implementation. We present microfluidic and optical networks as a basis for the interconnection of optofluidic sensor modules. Finally, we show the potential for entire optofluidic networks.

  16. Modular Platforms for Optofluidic Systems

    NASA Astrophysics Data System (ADS)

    Brammer, Marko; Mappes, Timo

    2014-01-01

    Optofluidics is increasingly gaining impact in a number of different fields of research, namely biology and medicine, environmental monitoring and green energy. However, the market for optofluidic products is still in the early development phase. In this manuscript, we discuss modular platforms as a potential concept to facilitate the transfer of optofluidic sensing systems to an industrial implementation. We present microfluidic and optical networks as a basis for the interconnection of optofluidic sensor modules. Finally, we show the potential for entire optofluidic networks

  17. Multidimensional bioseparation with modular microfluidics

    DOEpatents

    Chirica, Gabriela S.; Renzi, Ronald F.

    2013-08-27

    A multidimensional chemical separation and analysis system is described including a prototyping platform and modular microfluidic components capable of rapid and convenient assembly, alteration and disassembly of numerous candidate separation systems. Partial or total computer control of the separation system is possible. Single or multiple alternative processing trains can be tested, optimized and/or run in parallel. Examples related to the separation and analysis of human bodily fluids are given.

  18. CAMAC modular programmable function generator

    SciTech Connect

    Turner, G.W.; Suehiro, S.; Hendricks, R.W.

    1980-12-01

    A CAMAC modular programmable function generator has been developed. The device contains a 1024 word by 12-bit memory, a 12-bit digital-to-analog converter with a 600 ns settling time, an 18-bit programmable frequency register, and two programmable trigger output registers. The trigger registers can produce programmed output logic transitions at various (binary) points in the output function curve, and are used to synchronize various other data acquisition devices with the function curve.

  19. Estimation of inter-modular connectivity from the local field potentials in a hierarchical modular network

    NASA Astrophysics Data System (ADS)

    Cui, Xue-Mei; Kim, Won Sup; Hwang, Dong-Uk; Han, Seung Kee

    2015-05-01

    We propose a method of estimating inter-modular connectivity in a hierarchical modular network. The method is based on an analysis of inverse phase synchronization applied to the local field potentials on a hierarchical modular network of phase oscillators. For a strong-coupling strength, the inverse phase synchronization index of the local field potentials for two modules depends linearly on the corresponding inter-modular connectivity defined as the number of links connecting the modules. The method might enable us to estimate the inter-modular connectivity in various complex systems from the inverse phase synchronization index of the mesoscopic modular activities.

  20. Inhibition of Grape Crown Gall by Agrobacterium vitis F2/5 Requires Two Nonribosomal Peptide Synthetases and One Polyketide Synthase.

    PubMed

    Zheng, Desen; Burr, Thomas J

    2016-02-01

    Agrobacterium vitis nontumorigenic strain F2/5 is able to inhibit crown gall disease on grapevines. The mechanism of grape tumor inhibition (GTI) by F2/5 has not been fully determined. In this study, we demonstrate that two nonribosomal peptide synthetase (NRPS) genes (F-avi3342 and F-avi5730) and one polyketide synthase gene (F-avi4330) are required for GTI. Knockout of any one of them resulted in F/25 losing GTI capacity. We previously reported that F-avi3342 and F-avi4330 but not F-avi5730 are required for induction of grape tissue necrosis and tobacco hypersensitive response. F-avi5730 is predicted to encode a single modular NRPS. It is located in a cluster that is homologous to the siderophore vicibactin biosynthesis locus in Rhizobium species. Individual disruption of F-avi5730 and two immediate downstream genes, F-avi5731 and F-avi5732, all resulted in reduced siderophore production; however, only F-avi5730 was found to be required for GTI. Complemented F-avi5730 mutant (ΔF-avi5730(+)) restored a wild-type level of GTI activity. It was determined that, over time, populations of ΔF-avi4330, ΔF-avi3342, and ΔF-avi5730 at inoculated wound sites on grapevine did not differ from those of ΔF-avi5730(+) indicating that loss of GTI was not due to reduced colonization of wound sites by mutants.

  1. Cyclic modular beta-sheets.

    PubMed

    Woods, R Jeremy; Brower, Justin O; Castellanos, Elena; Hashemzadeh, Mehrnoosh; Khakshoor, Omid; Russu, Wade A; Nowick, James S

    2007-03-07

    The development of peptide beta-hairpins is problematic, because folding depends on the amino acid sequence and changes to the sequence can significantly decrease folding. Robust beta-hairpins that can tolerate such changes are attractive tools for studying interactions involving protein beta-sheets and developing inhibitors of these interactions. This paper introduces a new class of peptide models of protein beta-sheets that addresses the problem of separating folding from the sequence. These model beta-sheets are macrocyclic peptides that fold in water to present a pentapeptide beta-strand along one edge; the other edge contains the tripeptide beta-strand mimic Hao [JACS 2000, 122, 7654] and two additional amino acids. The pentapeptide and Hao-containing peptide strands are connected by two delta-linked ornithine (deltaOrn) turns [JACS 2003, 125, 876]. Each deltaOrn turn contains a free alpha-amino group that permits the linking of individual modules to form divalent beta-sheets. These "cyclic modular beta-sheets" are synthesized by standard solid-phase peptide synthesis of a linear precursor followed by solution-phase cyclization. Eight cyclic modular beta-sheets 1a-1h containing sequences based on beta-amyloid and macrophage inflammatory protein 2 were synthesized and characterized by 1H NMR. Linked cyclic modular beta-sheet 2, which contains two modules of 1b, was also synthesized and characterized. 1H NMR studies show downfield alpha-proton chemical shifts, deltaOrn delta-proton magnetic anisotropy, and NOE cross-peaks that establish all compounds but 1c and 1g to be moderately or well folded into a conformation that resembles a beta-sheet. Pulsed-field gradient NMR diffusion experiments show little or no self-association at low (

  2. Genome mining: Prediction of lipopeptides and polyketides from Bacillus and related Firmicutes

    PubMed Central

    Aleti, Gajender; Sessitsch, Angela; Brader, Günter

    2015-01-01

    Bacillus and related genera in the Bacillales within the Firmicutes harbor a variety of secondary metabolite gene clusters encoding polyketide synthases and non-ribosomal peptide synthetases responsible for remarkable diverse number of polyketides (PKs) and lipopeptides (LPs). These compounds may be utilized for medical and agricultural applications. Here, we summarize the knowledge on structural diversity and underlying gene clusters of LPs and PKs in the Bacillales. Moreover, we evaluate by using published prediction tools the potential metabolic capacity of these bacteria to produce type I PKs or LPs. The huge sequence repository of bacterial genomes and metagenomes provides the basis for such genome-mining to reveal the potential for novel structurally diverse secondary metabolites. The otherwise cumbersome task to isolate often unstable PKs and deduce their structure can be streamlined. Using web based prediction tools, we identified here several novel clusters of PKs and LPs from genomes deposited in the database. Our analysis suggests that a substantial fraction of predicted LPs and type I PKs are uncharacterized, and their functions remain to be studied. Known and predicted LPs and PKs occurred in the majority of the plant associated genera, predominantly in Bacillus and Paenibacillus. Surprisingly, many genera from other environments contain no or few of such compounds indicating the role of these secondary metabolites in plant-associated niches. PMID:25893081

  3. A conserved motif flags Acyl Carrier Proteins for β-branching in polyketide synthesis

    PubMed Central

    Song, Zhongshu; Farmer, Rohit; Williams, Christopher; Hothersall, Joanne; Płoskoń, Eliza; Wattana-amorn, Pakorn; Stephens, Elton R.; Yamada, Erika; Gurney, Rachel; Takebayashi, Yuiko; Masschelein, Joleen; Cox, Russell J.; Lavigne, Rob; Willis, Christine L.; Simpson, Thomas J.; Crosby, John; Winn, Peter J.; Thomas, Christopher M.; Crump, Matthew P.

    2015-01-01

    Type I PKSs often utilise programmed β-branching, via enzymes of an “HMG-CoA synthase (HCS) cassette”, to incorporate various side chains at the second carbon from the terminal carboxylic acid of growing polyketide backbones. We identified a strong sequence motif in Acyl Carrier Proteins (ACPs) where β-branching is known. Substituting ACPs confirmed a correlation of ACP type with β-branching specificity. While these ACPs often occur in tandem, NMR analysis of tandem β-branching ACPs indicated no ACP-ACP synergistic effects and revealed that the conserved sequence motif forms an internal core rather than an exposed patch. Modelling and mutagenesis identified ACP Helix III as a probable anchor point of the ACP-HCS complex whose position is determined by the core. Mutating the core affects ACP functionality while ACP-HCS interface substitutions modulate system specificity. Our method for predicting β-carbon branching expands the potential for engineering novel polyketides and lays a basis for determining specificity rules. PMID:24056399

  4. Detection of polyketide synthase and nonribosomal peptide synthetase biosynthetic genes from antimicrobial coral-associated actinomycetes.

    PubMed

    Li, Jie; Dong, Jun-De; Yang, Jian; Luo, Xiong-Ming; Zhang, Si

    2014-10-01

    The diversity and properties of actinobacteria, predominant residents in coral holobionts, have been rarely documented. In this study, we aimed to explore the species diversity, antimicrobial activities and biosynthetic potential of culturable actinomycetes within the tissues of the scleractinian corals Porites lutea, Galaxea fascicularis and Acropora millepora from the South China Sea. A total of 70 strains representing 13 families and 15 genera of actinobacteria were isolated. The antimicrobial activity and biosynthetic potential of fifteen representative filamentous actinomycetes were estimated. Crude fermentation extracts of 6 strains exhibited comparable or greater activities against Vibrio alginolyticus than ciprofloxacin. Seven of the 15 actinomycetes strains possess type I polyketide synthases (PKS-I) and/or nonribosomal peptide synthetases (NRPS) genes. Nine tested strains possess type II polyketide synthases (PKS-II). Phylogenetic analysis based on 16S rRNA gene sequences indicated that these PKS and NRPS gene screening positive strains belong to genera Nocardiopsis, Pseudonocardia, Streptomyces, Micromonospora, Amycolatopsis and Prauserella. One PKS-I and four NRPS fragments showed <70% similarity to their closest relatives, which suggested the novelty of these genes. This study helps uncover the genetic capacity of stony coral-associated actinomycetes to produce bioactive molecules.

  5. Type III polyketide synthase is involved in the biosynthesis of protocatechuic acid in Aspergillus niger.

    PubMed

    Lv, Yangyong; Xiao, Jing; Pan, Li

    2014-11-01

    Genomic studies have shown that not only plants but also filamentous fungi contain type III polyketide synthases. To study the function of type III polyketide synthase (AnPKSIII) in Aspergillus niger, a deletion strain (delAnPKSIII) and an overexpression strain (oeAnPKSIII) were constructed in A. niger MA169.4, a derivative of the wild-type (WT) A. niger ATCC 9029 that produces large quantities of gluconic acid. Alterations in the metabolites were analyzed by HPLC when the extract of the overexpression strain was compared with extracts of the WT and deletion strains. Protocatechuic acid (PCA; 3,4-dihydroxybenzoic acid, 3.2 mg/l) was isolated and identified as the main product of AnPKSIII when inductively expressed in A. niger MA169.4. The molecular weight of PCA was 154.1 (m/z 153.1 [M-H](-)), was detected by ESI-MS in the negative ionization mode, and (1)H and (13)C NMR data confirmed its structure.

  6. Putative Monofunctional Type I Polyketide Synthase Units: A Dinoflagellate-Specific Feature?

    PubMed Central

    Eichholz, Karsten; Beszteri, Bánk; John, Uwe

    2012-01-01

    Marine dinoflagellates (alveolata) are microalgae of which some cause harmful algal blooms and produce a broad variety of most likely polyketide synthesis derived phycotoxins. Recently, novel polyketide synthesase (PKS) transcripts have been described from the Florida red tide dinoflagellate Karenia brevis (gymnodiniales) which are evolutionarily related to Type I PKS but were apparently expressed as monofunctional proteins, a feature typical of Type II PKS. Here, we investigated expression units of PKS I-like sequences in Alexandrium ostenfeldii (gonyaulacales) and Heterocapsa triquetra (peridiniales) at the transcript and protein level. The five full length transcripts we obtained were all characterized by polyadenylation, a 3′ UTR and the dinoflagellate specific spliced leader sequence at the 5′end. Each of the five transcripts encoded a single ketoacylsynthase (KS) domain showing high similarity to K. brevis KS sequences. The monofunctional structure was also confirmed using dinoflagellate specific KS antibodies in Western Blots. In a maximum likelihood phylogenetic analysis of KS domains from diverse PKSs, dinoflagellate KSs formed a clade placed well within the protist Type I PKS clade between apicomplexa, haptophytes and chlorophytes. These findings indicate that the atypical PKS I structure, i.e., expression as putative monofunctional units, might be a dinoflagellate specific feature. In addition, the sequenced transcripts harbored a previously unknown, apparently dinoflagellate specific conserved N-terminal domain. We discuss the implications of this novel region with regard to the putative monofunctional organization of Type I PKS in dinoflagellates. PMID:23139807

  7. Identification of a Polyketide Synthase Involved in Sorbicillin Biosynthesis by Penicillium chrysogenum

    PubMed Central

    Salo, Oleksandr; Guzmán-Chávez, Fernando; Ries, Marco I.; Lankhorst, Peter P.; Bovenberg, Roel A. L.; Vreeken, Rob J.

    2016-01-01

    ABSTRACT Secondary metabolism in Penicillium chrysogenum was intensively subjected to classical strain improvement (CSI), the resulting industrial strains producing high levels of β-lactams. During this process, the production of yellow pigments, including sorbicillinoids, was eliminated as part of a strategy to enable the rapid purification of β-lactams. Here we report the identification of the polyketide synthase (PKS) gene essential for sorbicillinoid biosynthesis in P. chrysogenum. We demonstrate that the production of polyketide precursors like sorbicillinol and dihydrosorbicillinol as well as their derivatives bisorbicillinoids requires the function of a highly reducing PKS encoded by the gene Pc21g05080 (pks13). This gene belongs to the cluster that was mutated and transcriptionally silenced during the strain improvement program. Using an improved β-lactam-producing strain, repair of the mutation in pks13 led to the restoration of sorbicillinoid production. This now enables genetic studies on the mechanism of sorbicillinoid biosynthesis in P. chrysogenum and opens new perspectives for pathway engineering. IMPORTANCE Sorbicillinoids are secondary metabolites with antiviral, anti-inflammatory, and antimicrobial activities produced by filamentous fungi. This study identified the gene cluster responsible for sorbicillinoid formation in Penicillium chrysogenum, which now allows engineering of this diverse group of compounds. PMID:27107123

  8. Koningiopisins A-H, Polyketides with Synergistic Antifungal Activities from the Endophytic Fungus Trichoderma koningiopsis.

    PubMed

    Liu, Kai; Yang, Yabin; Miao, Cui-Ping; Zheng, You-Kun; Chen, Jin-Lian; Chen, You-Wei; Xu, Li-Hua; Guang, Hui-Lin; Ding, Zhong-Tao; Zhao, Li-Xing

    2016-03-01

    Eight new fungal polyketides named koningiopisins A-H (1-8) and four previously known polyketides (9-12) were isolated from the endophytic fungus Trichoderma koningiopsis YIM PH 30002. Their structures were elucidated using extensive spectral data interpretation, and their antifungal and synergistic activities were also evaluated. Koningiopisin C (3) exhibited in vitro antifungal activity against the phytopathogenic fungus Plectosphaerella cucumerina with an MIC of 16 µg/mL. Although the antifungal activities of single compounds were not obvious, a mixture of six compounds (4-9) exhibited potent synergistic antifungal activity against P. cucumerina with an MIC of 16 µg/mL, and the antifungal activity of the mixture of any two compounds with a 1:1 ratio was better than that observed from the individual compound. The synergistic biological activity of the metabolites in YIM PH 30002 demonstrates the significant ecological function of the endophyte for its host plant, and provides additional insight into the search for and development of agents for biological control. Georg Thieme Verlag KG Stuttgart · New York.

  9. Production of the polyketide 6-MSA in yeast engineered for increased malonyl-CoA supply.

    PubMed

    Wattanachaisaereekul, Songsak; Lantz, Anna Eliasson; Nielsen, Michael Lynge; Nielsen, Jens

    2008-09-01

    The heterologous production of fungal polyketides was investigated using 6-methylsalicylic acid synthase (6-MSAS) as a model polyketide synthase and Saccharomyces cerevisiae as a host. In order to improve the production of 6-MSA by enhancing the supply of precursors, the promoter of the gene (ACC1) encoding acetyl-CoA carboxylase, which catalyzes the conversion of acetyl-CoA to malonyl-CoA, was replaced with a strong, constitutive promoter (TEF1p) in a strain harboring two plasmids carrying the genes encoding 6-MSAS from Penicillium patulum and PPTase from Aspergillus nidulans, respectively. The strain was characterized in batch cultivations with a glucose minimal media (20 g/L), and a 60% increase in 6-MSA titer was observed compared to a strain having the native promoter in front of ACC1. The production of 6-MSA was scaled up by the cultivation in minimal media containing 50 g/L of glucose, and hereby a final titer of 554+/-26 mg/L of 6-MSA was obtained.

  10. Biosynthesis of antifungal and antibacterial polyketides by Burkholderia gladioli in coculture with Rhizopus microsporus.

    PubMed

    Ross, Claudia; Opel, Viktoria; Scherlach, Kirstin; Hertweck, Christian

    2014-12-01

    Fungi-bacteria interactions can impact the course of fungal infection and biotechnological use. The mucoralean fungus Rhizopus microsporus, traditionally used in food fermentations (tempe and sufu), is frequently accompanied by Burkholderia gladioli pv. cocovenenans. When producing tempe bongkrek, the bacterial contamination can lead to lethal food-related intoxications caused by the respiratory toxin bongkrekic acid. To unveil the metabolic potential of the fungus-associated bacterium, we sequenced its genome, assigned secondary metabolite biosynthesis gene clusters and monitored the metabolic profile under various growth conditions. In addition to the bongkrekic acid biosynthesis gene cluster we found gene clusters coding for the biosynthesis of toxoflavin and a complex polyketide. The orphan polyketide synthase gene cluster was activated under conditions that emulate tempe production, which enabled isolation and structure elucidation of four members of the enacyloxin family of antibiotics, out of which one is new. Moreover, we found that the fungus positively influences the growth of the bacteria and dramatically increases bongkrekic acid production in stationary culture, which inhibits the growth of the fungus. These results showcase the context-dependent formation of antifungal and antibacterial agents at the fungal-bacterial interface, which may also serve as a model for scenarios observed in mixed infections. © 2014 Blackwell Verlag GmbH.

  11. Recent advancements in the biosynthetic mechanisms for polyketide-derived mycotoxins.

    PubMed

    Huffman, Justin; Gerber, Ryan; Du, Liangcheng

    2010-09-01

    Polyketides (PKs) are a large group of natural products produced by microorganisms and plants. They are biopolymers of acetate and other short carboxylates and are biosynthesized by multifunctional enzymes called polyketide synthases (PKSs). This review discusses the biosynthesis of four toxic PK, aflatoxins, fumonisins, ochratoxins (OTs), and zearalenone. These metabolites are structurally diverse and differ in their mechanisms of toxicity. However, they are all of concern in food safety and agriculture because of their toxic properties and their frequent accumulation in crops used for food and feed. The focus is on the recent advancements in the understanding of the molecular mechanisms for the biosynthesis of these mycotoxins. Several of the mycotoxin PKSs have been genetically and biochemically studied while other PKSs remain to be investigated. Multiple post-PKS modifications are often required for the maturation of the mycotoxins. Many of these modification steps for aflatoxins and fumonisins are well established while the post-PKS modifications for zearalenone and OTs remain to be biochemically characterized. More efforts are needed to completely illustrate the biosynthetic mechanisms for this important group of PKs. Copyright 2010 Wiley Periodicals, Inc.

  12. A proteomic survey of nonribosomal peptide and polyketide biosynthesis in actinobacteria.

    PubMed

    Chen, Yunqiu; Ntai, Ioanna; Ju, Kou-San; Unger, Michelle; Zamdborg, Leonid; Robinson, Sarah J; Doroghazi, James R; Labeda, David P; Metcalf, William W; Kelleher, Neil L

    2012-01-01

    Actinobacteria such as streptomycetes are renowned for their ability to produce bioactive natural products including nonribosomal peptides (NRPs) and polyketides (PKs). The advent of genome sequencing has revealed an even larger genetic repertoire for secondary metabolism with most of the small molecule products of these gene clusters still unknown. Here, we employed a "protein-first" method called PrISM (Proteomic Investigation of Secondary Metabolism) to screen 26 unsequenced actinomycetes using mass spectrometry-based proteomics for the targeted detection of expressed nonribosomal peptide synthetases or polyketide synthases. Improvements to the original PrISM screening approach (Nat. Biotechnol. 2009, 27, 951-956), for example, improved de novo peptide sequencing, have enabled the discovery of 10 NRPS/PKS gene clusters from 6 strains. Taking advantage of the concurrence of biosynthetic enzymes and the secondary metabolites they generate, two natural products were associated with their previously "orphan" gene clusters. This work has demonstrated the feasibility of a proteomics-based strategy for use in screening for NRP/PK production in actinomycetes (often >8 Mbp, high GC genomes) versus the bacilli (2-4 Mbp genomes) used previously.

  13. Exploring the chemistry of uncultivated bacterial symbionts: antitumor polyketides of the pederin family.

    PubMed

    Piel, Jörn; Butzke, Daniel; Fusetani, Nobuhiro; Hui, Dequan; Platzer, Matthias; Wen, Gaiping; Matsunaga, Shigeki

    2005-03-01

    Symbiotic bacteria have long been proposed as being responsible for the production of numerous natural products isolated from invertebrate animals. However, systematic studies of invertebrate-symbiont associations are usually associated with serious technical challenges, such as the general resistance of symbionts to culturing attempts and the complexity of many microbial consortia. Herein an overview is provided on the culture-independent, metagenomic strategies recently employed by our group to contribute to a better understanding of natural product symbiosis. Using terrestrial Paederus spp. beetles and the marine sponge Theonella swinhoei as model animals, the putative genes responsible for the production of pederin-type antitumor polyketides have been isolated. In Paederus fuscipes, which uses pederin for chemical defense, these genes belong to an as-yet unculturable symbiont closely related to Pseudomonas aeruginosa. To study the extremely complex association of T. swinhoei and its multispecies bacterial consortium, we used a phylogenetic approach that allowed the isolation of onnamide/theopederin polyketide synthase genes from an uncultured sponge symbiont. Analysis of the biosynthesis genes provided unexpected insights into a possible evolution of pederin-type pathways. Besides revealing new facets of invertebrate chemical ecology, these first gene clusters from uncultivated symbiotic producers suggest possible biotechnological strategies to solve the supply problem associated with the development of most marine drug candidates.

  14. Total Biosynthesis and Diverse Applications of the Nonribosomal Peptide-Polyketide Siderophore Yersiniabactin

    PubMed Central

    Ahmadi, Mahmoud Kamal; Fawaz, Samar; Jones, Charles H.; Zhang, Guojian

    2015-01-01

    Yersiniabactin (Ybt) is a mixed nonribosomal peptide-polyketide natural product natively produced by the pathogen Yersinia pestis. The compound enables iron scavenging capabilities upon host infection and is biosynthesized by a nonribosomal peptide synthetase featuring a polyketide synthase module. This pathway has been engineered for expression and biosynthesis using Escherichia coli as a heterologous host. In the current work, the biosynthetic process for Ybt formation was improved through the incorporation of a dedicated step to eliminate the need for exogenous salicylate provision. When this improvement was made, the compound was tested in parallel applications that highlight the metal-chelating nature of the compound. In the first application, Ybt was assessed as a rust remover, demonstrating a capacity of ∼40% compared to a commercial removal agent and ∼20% relative to total removal capacity. The second application tested Ybt in removing copper from a variety of nonbiological and biological solution mixtures. Success across a variety of media indicates potential utility in diverse scenarios that include environmental and biomedical settings. PMID:26025901

  15. The Polyketide Synthase Gene pks4 of Trichoderma reesei Provides Pigmentation and Stress Resistance

    PubMed Central

    Atanasova, Lea; Knox, Benjamin P.; Kubicek, Christian P.; Baker, Scott E.

    2013-01-01

    Species of the fungal genus Trichoderma (Hypocreales, Ascomycota) are well-known for their production of various secondary metabolites. Nonribosomal peptides and polyketides represent a major portion of these products. In a recent phylogenomic investigation of Trichoderma polyketide synthase (PKS)-encoding genes, the pks4 from T. reesei was shown to be an orthologue of pigment-forming PKSs involved in synthesis of aurofusarin and bikaverin in Fusarium spp. In this study, we show that deletion of this gene in T. reesei results in loss of green conidial pigmentation and in pigmentation alteration of teleomorph structures. It also has an impact on conidial cell wall stability and the antagonistic abilities of T. reesei against other fungi, including formation of inhibitory metabolites. In addition, deletion of pks4 significantly influences the expression of other PKS-encoding genes of T. reesei. To our knowledge, this is the first indication that a low-molecular-weight pigment-forming PKS is involved in defense, mechanical stability, and stress resistance in fungi. PMID:24036343

  16. Redundant synthesis of a conidial polyketide by two distinct secondary metabolite clusters in Aspergillus fumigatus

    PubMed Central

    Throckmorton, Kurt; Lim, Fang Yun; Kontoyiannis, Dimitrios P.; Zheng, Weifa; Keller, Nancy P.

    2016-01-01

    Summary Filamentous fungi are renowned for the production of bioactive secondary metabolites. Typically, one distinct metabolite is generated from a specific secondary metabolite cluster. Here, we characterize the newly described trypacidin (tpc) cluster in the opportunistic human pathogen Aspergillus fumigatus. We find that this cluster as well as the previously characterized endocrocin (enc) cluster both contribute to the production of the spore metabolite endocrocin. Whereas trypacidin is eliminated when only tpc cluster genes are deleted, endocrocin production is only eliminated when both the tpc and enc non-reducing polyketide synthase-encoding genes, tpcC and encA, respectively, are deleted. EncC, an anthrone oxidase, converts the product released from EncA to endocrocin as a final product. In contrast, endocrocin synthesis by the tpc cluster likely results from incomplete catalysis by TpcK (a putative decarboxylase), as its deletion results in a nearly 10-fold increase in endocrocin production. We suggest endocrocin is likely a shunt product in all related non-reducing polyketide synthase clusters containing homologues of TpcK and TpcL (a putative anthrone oxidase), e.g. geodin and monodictyphenone. This finding represents an unusual example of two physically discrete secondary metabolite clusters generating the same natural product in one fungal species by distinct routes. PMID:26242966

  17. Engineering a Polyketide Synthase for In Vitro Production of Adipic Acid.

    PubMed

    Hagen, Andrew; Poust, Sean; Rond, Tristan de; Fortman, Jeffrey L; Katz, Leonard; Petzold, Christopher J; Keasling, Jay D

    2016-01-15

    Polyketides have enormous structural diversity, yet polyketide synthases (PKSs) have thus far been engineered to produce only drug candidates or derivatives thereof. Thousands of other molecules, including commodity and specialty chemicals, could be synthesized using PKSs if composing hybrid PKSs from well-characterized parts derived from natural PKSs was more efficient. Here, using modern mass spectrometry techniques as an essential part of the design-build-test cycle, we engineered a chimeric PKS to enable production one of the most widely used commodity chemicals, adipic acid. To accomplish this, we introduced heterologous reductive domains from various PKS clusters into the borrelidin PKS' first extension module, which we previously showed produces a 3-hydroxy-adipoyl intermediate when coincubated with the loading module and a succinyl-CoA starter unit. Acyl-ACP intermediate analysis revealed an unexpected bottleneck at the dehydration step, which was overcome by introduction of a carboxyacyl-processing dehydratase domain. Appending a thioesterase to the hybrid PKS enabled the production of free adipic acid. Using acyl-intermediate based techniques to "debug" PKSs as described here, it should one day be possible to engineer chimeric PKSs to produce a variety of existing commodity and specialty chemicals, as well as thousands of chemicals that are difficult to produce from petroleum feedstocks using traditional synthetic chemistry.

  18. Minimal Streptomyces sp. strain C5 daunorubicin polyketide biosynthesis genes required for aklanonic acid biosynthesis.

    PubMed Central

    Rajgarhia, V B; Strohl, W R

    1997-01-01

    The structure of the Streptomyces sp. strain C5 daunorubicin type II polyketide synthase (PKS) gene region is different from that of other known type II PKS gene clusters. Directly downstream of the genes encoding ketoacylsynthase alpha and beta (KS alpha, KS beta) are two genes (dpsC, dpsD) encoding proteins of unproven function, both absent from other type II PKS gene clusters. Also in contrast to other type II PKS clusters, the gene encoding the acyl carrier protein (ACP), dpsG, is located about 6.8 kbp upstream of the genes encoding the daunorubicin KS alpha and KS beta. In this work, we demonstrate that the minimal genes required to produce aklanonic acid in heterologous hosts are dpsG (ACP), dauI (regulatory activator), dpsA (KS alpha), dpsB (KS beta), dpsF (aromatase), dpsE (polyketide reductase), and dauG (putative deoxyaklanonic acid oxygenase). The two unusual open reading frames, dpsC (KASIII homolog lacking a known active site) and dpsD (acyltransferase homolog), are not required to synthesize aklanonic acid. Additionally, replacement of dpsD or dpsCD in Streptomyces sp. strain C5 with a neomycin resistance gene (aphI) results in mutant strains that still produced anthracyclines. PMID:9098068

  19. Lindgomycin, an Unusual Antibiotic Polyketide from a Marine Fungus of the Lindgomycetaceae

    PubMed Central

    Wu, Bin; Wiese, Jutta; Labes, Antje; Kramer, Annemarie; Schmaljohann, Rolf; Imhoff, Johannes F.

    2015-01-01

    An unusual polyketide with a new carbon skeleton, lindgomycin (1), and the recently described ascosetin (2) were extracted from mycelia and culture broth of different Lindgomycetaceae strains, which were isolated from a sponge of the Kiel Fjord in the Baltic Sea (Germany) and from the Antarctic. Their structures were established by spectroscopic means. In the new polyketide, two distinct domains, a bicyclic hydrocarbon and a tetramic acid, are connected by a bridging carbonyl. The tetramic acid substructure of compound 1 was proved to possess a unique 5-benzylpyrrolidine-2,4-dione unit. The combination of 5-benzylpyrrolidine-2,4-dione of compound 1 in its tetramic acid half and 3-methylbut-3-enoic acid pendant in its decalin half allow the assignment of a new carbon skeleton. The new compound 1 and ascosetin showed antibiotic activities with IC50 value of 5.1 (±0.2) µM and 3.2 (±0.4) μM, respectively, against methicillin-resistant Staphylococcus aureus. PMID:26225984

  20. A type III polyketide synthase from Wachendorfia thyrsiflora and its role in diarylheptanoid and phenylphenalenone biosynthesis.

    PubMed

    Brand, S; Hölscher, D; Schierhorn, A; Svatos, A; Schröder, J; Schneider, B

    2006-07-01

    Chalcone synthase (CHS) related type III plant polyketide synthases (PKSs) are likely to be involved in the biosynthesis of diarylheptanoids (e.g. curcumin and polycyclic phenylphenalenones), but no such activity has been reported. Root cultures from Wachendorfia thyrsiflora (Haemodoraceae) are a suitable source to search for such enzymes because they synthesize large amounts of phenylphenalenones, but no other products that are known to require CHSs or related enzymes (e.g. flavonoids or stilbenes). A homology-based RT-PCR strategy led to the identification of cDNAs for a type III PKS sharing only approximately 60% identity with typical CHSs. It was named WtPKS1 (W. thyrsiflora polyketide synthase 1). The purified recombinant protein accepted a large variety of aromatic and aliphatic starter CoA esters, including phenylpropionyl- and side-chain unsaturated phenylpropanoid-CoAs. The simplest model for the initial reaction in diarylheptanoid biosynthesis predicts a phenylpropanoid-CoA as starter and a single condensation reaction to a diketide. Benzalacetones, the expected release products, were observed only with unsaturated phenylpropanoid-CoAs, and the best results were obtained with 4-coumaroyl-CoA (80% of the products). With all other substrates, WtPKS1 performed two condensation reactions and released pyrones. We propose that WtPKS1 catalyses the first step in diarylheptanoid biosynthesis and that the observed pyrones are derailment products in the absence of downstream processing proteins.

  1. Parameterization of OPLS-AA force field for the conformational analysis of macrocyclic polyketides.

    PubMed

    Kahn, Kalju; Bruice, Thomas C

    2002-07-30

    The parameters for the OPLS-AA potential energy function have been extended to include some functional groups that are present in macrocyclic polyketides. Existing OPLS-AA torsional parameters for alkanes, alcohols, ethers, hemiacetals, esters, and ketoamides were improved based on MP2/aug-cc-pVTZ and MP2/aug-cc-pVDZ calculations. Nonbonded parameters for the sp(3) carbon and oxygen atoms were refined using Monte Carlo simulations of bulk liquids. The resulting force field predicts conformer energies and torsional barriers of alkanes, alcohols, ethers, and hemiacetals with an overall RMS deviation of 0.40 kcal/mol as compared to reference data. Densities of 19 bulk liquids are predicted with an average error of 1.1%, and heats of vaporization are reproduced within 2.4% of experimental values. The force field was used to perform conformational analysis of smaller analogs of the macrocyclic polyketide drug FK506. Structures that adopted low-energy conformations similar to that of bound FK506 were identified. The results show that a linker of four ketide units constitutes the shortest effector domain that allows binding of the ketide drugs to FKBP proteins. It is proposed that the exact chemical makeup of the effector domain has little influence on the conformational preference of tetraketides.

  2. [Diversity of polyketide synthase genes (PKS) in metagenomic community of the freshwater sponge].

    PubMed

    Kaliuzhnaia, O V; Kulakova, N V; Itskovich, B V

    2012-01-01

    Screening of metagenomic DNA of microbial community, associated with Baikalian sponge Lubomirskia baicalensis, was made to show the presence of polyketide synthase genes (PKS). PKS enzymatic systems take part in synthesis of a great number of biologically-active substances. Cloning and sequencing of amplified products of the ketosynthase domain section of PKS gene cluster has revealed 15 fragments of PKS genes differing from each other's on 35-65% by aminoacid sequences. BLASTX analysis has shown that all these sequences belong to the KS-domains identified in various groups of microorganisms: alpha-, beta-, delta-Proteobacteria, Verrucomicrobia, Cyanobacteria, Chlorophyta. Some sequences were related to the genes which are taking part in biosynthesis of curacin A (CurI, CurJ), stigmatellin (StiC, StiG), nostophycin (NpnB), and cryptophycins (CrpB). The homology of the found sequences with those of the EMBL database varies within 50-82% confirming the presence in fresh-water sponge community the genes for synthesis of the new, yet not studied polyketide substances, possessing the biotechnological potential.

  3. A Proteomic Survey of Nonribosomal Peptide and Polyketide Biosynthesis in Actinobacteria

    PubMed Central

    Chen, Yunqiu; Ntai, Ioanna; Ju, Kou-San; Unger, Michelle; Zamdborg, Leonid; Robinson, Sarah J.; Doroghazi, James R.; Labeda, David P.; Metcalf, William W.; Kelleher, Neil L.

    2011-01-01

    Actinobacteria such as streptomycetes are renowned for their ability to produce bioactive natural products including nonribosomal peptides (NRPs) and polyketides (PKs). The advent of genome sequencing has revealed an even larger genetic repertoire for secondary metabolism with most of the small molecule products of these gene clusters still unknown. Here, we employed a “protein-first” method called PrISM (Proteomic Investigation of Secondary Metabolism) to screen 26 unsequenced actinomycetes using mass spectrometry-based proteomics for the targeted detection of expressed nonribosomal peptide synthetases or polyketide synthases. Improvements to the original PrISM screening approach (Nature Biotechnology, 2009, 27, 951 – 956), e.g. improved de novo peptide sequencing, have enabled the discovery of ten NRPS/PKS gene clusters from six strains. Taking advantage of the concurrence of biosynthetic enzymes and the secondary metabolites they generate, two natural products were associated with their previously ‘orphan’ gene clusters. This work has demonstrated the feasibility of a proteomics-based strategy for use in screening for NRP/PK production in actinomycetes (often >8 Mbp, high GC genomes) versus the bacilli (2–4 Mbp genomes) used previously. PMID:21978092

  4. Engineering a Polyketide Synthase for In Vitro Production of Adipic Acid

    SciTech Connect

    Hagen, Andrew; Poust, Sean; Rond, Tristan de; Fortman, Jeffrey L.; Katz, Leonard; Petzold, Christopher J.; Keasling, Jay D.

    2015-10-26

    Polyketides have enormous structural diversity, yet polyketide synthases (PKSs) have thus far been engineered to produce only drug candidates or derivatives thereof. Thousands of other molecules, including commodity and specialty chemicals, could be synthesized using PKSs if composing hybrid PKSs from well-characterized parts derived from natural PKSs was more efficient. Here, using modern mass spectrometry techniques as an essential part of the design–build–test cycle, we engineered a chimeric PKS to enable production one of the most widely used commodity chemicals, adipic acid. To accomplish this, we introduced heterologous reductive domains from various PKS clusters into the borrelidin PKS’ first extension module, which we previously showed produces a 3-hydroxy-adipoyl intermediate when coincubated with the loading module and a succinyl-CoA starter unit. Acyl-ACP intermediate analysis revealed an unexpected bottleneck at the dehydration step, which was overcome by introduction of a carboxyacyl-processing dehydratase domain. Appending a thioesterase to the hybrid PKS enabled the production of free adipic acid. Using acyl-intermediate based techniques to “debug” PKSs as described here, it should one day be possible to engineer chimeric PKSs to produce a variety of existing commodity and specialty chemicals, as well as thousands of chemicals that are difficult to produce from petroleum feedstocks using traditional synthetic chemistry.

  5. Enhanced salinomycin production by adjusting the supply of polyketide extender units in Streptomyces albus.

    PubMed

    Lu, Chenyang; Zhang, Xiaojie; Jiang, Ming; Bai, Linquan

    2016-05-01

    The anticoccidial salinomycin is a polyketide produced by Streptomyces albus and requires malonyl-CoAs, methylmalonyl-CoAs, and ethylmalonyl-CoAs for the backbone assembly. Genome sequencing of S. albus DSM 41398 revealed a high percentage of genes involved in lipid metabolism, supporting the high salinomycin yield in oil-rich media. Seven PKS/PKS-NRPS gene clusters in the genome were found to be actively transcribed and had been individually deleted, which resulted in significantly improved salinomycin production. However, a combined deletion of PKS-NRPS-2 and PKS-6 showed no further improvement. Whereas the concentrations of malonyl-CoA and methylmalonyl-CoA were increased, the concentration of ethylmalonyl-CoA remained low in the mutants. An endogenous crotonyl-CoA reductase gene (ccr) was overexpressed in the ΔPKS-NRPS-2/ΔPKS-6 mutant, resulting in improved production. Combination of cluster deletions and over-expression of ccr gene led to an overall titer improvement of salinomycin from 0.60 to 6.60g/L. This engineering strategy can be implemented for various natural polyketides production.

  6. The polyketide synthase gene pks4 of Trichoderma reesei provides pigmentation and stress resistance.

    PubMed

    Atanasova, Lea; Knox, Benjamin P; Kubicek, Christian P; Druzhinina, Irina S; Baker, Scott E

    2013-11-01

    Species of the fungal genus Trichoderma (Hypocreales, Ascomycota) are well-known for their production of various secondary metabolites. Nonribosomal peptides and polyketides represent a major portion of these products. In a recent phylogenomic investigation of Trichoderma polyketide synthase (PKS)-encoding genes, the pks4 from T. reesei was shown to be an orthologue of pigment-forming PKSs involved in synthesis of aurofusarin and bikaverin in Fusarium spp. In this study, we show that deletion of this gene in T. reesei results in loss of green conidial pigmentation and in pigmentation alteration of teleomorph structures. It also has an impact on conidial cell wall stability and the antagonistic abilities of T. reesei against other fungi, including formation of inhibitory metabolites. In addition, deletion of pks4 significantly influences the expression of other PKS-encoding genes of T. reesei. To our knowledge, this is the first indication that a low-molecular-weight pigment-forming PKS is involved in defense, mechanical stability, and stress resistance in fungi.

  7. Modular microrobot for swimming in heterogeneous environments

    NASA Astrophysics Data System (ADS)

    Cheang, U. Kei; Meshkati, Meshkati; Fu, Henry; Kim, Minjun; Drexel University Team; University of Nevada, Reno Team

    2015-11-01

    One of the difficulties in navigating in vivo is to overcome many types of environments. This includes blood vessels of different diameters, fluids with different mechanical properties, and physical barriers. Inspired by conventional modular robotics, we demonstrate modular microrobotics using magnetic particles as the modular units to change size and shape through docking and undocking. Much like the vast variety of microorganisms navigating many different bio-environments, modular microswimmers have the ability to dynamically adapt different environments by reconfiguring the swimmers' physical characteristics. We model the docking as magnetic assembly and undocking mechanisms as deformation by hydrodynamic forces. We characterize the swimming capability of the modular microswimmer with different size and shapes. Finally, we demonstrate modular microrobotics by assembling a three-bead microswimmer into a nine-bead microswimmer, and then disassemble it into several independently swimming microswimmers..

  8. Modular Design in Treaty Verification Equipment

    SciTech Connect

    Macarthur, Duncan Whittemore; Benz, Jacob; Tolk, Keith; Weber, Tom

    2015-01-27

    It is widely believed that modular design is a good thing. However, there are often few explicit arguments, or even an agreed range of definitions, to back up this belief. In this paper, we examine the potential range of design modularity, the implications of various amounts of modularity, and the advantages and disadvantages of each level of modular construction. We conclude with a comparison of the advantages and disadvantages of each type, as well as discuss many caveats that should be observed to take advantage of the positive features of modularity and minimize the effects of the negative. The tradeoffs described in this paper will be evaluated during the conceptual design to determine what amount of modularity should be included.

  9. Modular workcells: modern methods for laboratory automation.

    PubMed

    Felder, R A

    1998-12-01

    Laboratory automation is beginning to become an indispensable survival tool for laboratories facing difficult market competition. However, estimates suggest that only 8% of laboratories will be able to afford total laboratory automation systems. Therefore, automation vendors have developed alternative hardware configurations called 'modular automation', to fit the smaller laboratory. Modular automation consists of consolidated analyzers, integrated analyzers, modular workcells, and pre- and post-analytical automation. These terms will be defined in this paper. Using a modular automation model, the automated core laboratory will become a site where laboratory data is evaluated by trained professionals to provide diagnostic information to practising physicians. Modem software information management and process control tools will complement modular hardware. Proper standardization that will allow vendor-independent modular configurations will assure success of this revolutionary new technology.

  10. Modular, Hierarchical Learning By Artificial Neural Networks

    NASA Technical Reports Server (NTRS)

    Baldi, Pierre F.; Toomarian, Nikzad

    1996-01-01

    Modular and hierarchical approach to supervised learning by artificial neural networks leads to neural networks more structured than neural networks in which all neurons fully interconnected. These networks utilize general feedforward flow of information and sparse recurrent connections to achieve dynamical effects. The modular organization, sparsity of modular units and connections, and fact that learning is much more circumscribed are all attractive features for designing neural-network hardware. Learning streamlined by imitating some aspects of biological neural networks.

  11. Modular, Hierarchical Learning By Artificial Neural Networks

    NASA Technical Reports Server (NTRS)

    Baldi, Pierre F.; Toomarian, Nikzad

    1996-01-01

    Modular and hierarchical approach to supervised learning by artificial neural networks leads to neural networks more structured than neural networks in which all neurons fully interconnected. These networks utilize general feedforward flow of information and sparse recurrent connections to achieve dynamical effects. The modular organization, sparsity of modular units and connections, and fact that learning is much more circumscribed are all attractive features for designing neural-network hardware. Learning streamlined by imitating some aspects of biological neural networks.

  12. Biosynthesis of the salinosporamide A polyketide synthase substrate chloroethylmalonyl-coenzyme A from S-adenosyl-l-methionine

    PubMed Central

    Eustáquio, Alessandra S.; McGlinchey, Ryan P.; Liu, Yuan; Hazzard, Christopher; Beer, Laura L.; Florova, Galina; Alhamadsheh, Mamoun M.; Lechner, Anna; Kale, Andrew J.; Kobayashi, Yoshihisa; Reynolds, Kevin A.; Moore, Bradley S.

    2009-01-01

    Polyketides are among the major classes of bioactive natural products used to treat microbial infections, cancer, and other diseases. Here we describe a pathway to chloroethylmalonyl-CoA as a polyketide synthase building block in the biosynthesis of salinosporamide A, a marine microbial metabolite whose chlorine atom is crucial for potent proteasome inhibition and anticancer activity. S-adenosyl-l-methionine (SAM) is converted to 5′-chloro-5′-deoxyadenosine (5′-ClDA) in a reaction catalyzed by a SAM-dependent chlorinase as previously reported. By using a combination of gene deletions, biochemical analyses, and chemical complementation experiments with putative intermediates, we now provide evidence that 5′-ClDA is converted to chloroethylmalonyl-CoA in a 7-step route via the penultimate intermediate 4-chlorocrotonyl-CoA. Because halogenation often increases the bioactivity of drugs, the availability of a halogenated polyketide building block may be useful in molecular engineering approaches toward polyketide scaffolds. PMID:19590008

  13. Alternative Sigma Factor Over-Expression Enables Heterologous Expression of a Type II Polyketide Biosynthetic Pathway in Escherichia coli

    PubMed Central

    Stevens, David Cole; Conway, Kyle R.; Pearce, Nelson; Villegas-Peñaranda, Luis Roberto; Garza, Anthony G.; Boddy, Christopher N.

    2013-01-01

    Background Heterologous expression of bacterial biosynthetic gene clusters is currently an indispensable tool for characterizing biosynthetic pathways. Development of an effective, general heterologous expression system that can be applied to bioprospecting from metagenomic DNA will enable the discovery of a wealth of new natural products. Methodology We have developed a new Escherichia coli-based heterologous expression system for polyketide biosynthetic gene clusters. We have demonstrated the over-expression of the alternative sigma factor σ54 directly and positively regulates heterologous expression of the oxytetracycline biosynthetic gene cluster in E. coli. Bioinformatics analysis indicates that σ54 promoters are present in nearly 70% of polyketide and non-ribosomal peptide biosynthetic pathways. Conclusions We have demonstrated a new mechanism for heterologous expression of the oxytetracycline polyketide biosynthetic pathway, where high-level pleiotropic sigma factors from the heterologous host directly and positively regulate transcription of the non-native biosynthetic gene cluster. Our bioinformatics analysis is consistent with the hypothesis that heterologous expression mediated by the alternative sigma factor σ54 may be a viable method for the production of additional polyketide products. PMID:23724102

  14. Nargenicin biosynthesis. Incorporation of polyketide chain elongation intermediates and support for a proposed intramolecular Diels-Alder cyclization

    SciTech Connect

    Cane, D.E.; Weitian Tan; Ott, W.R. )

    1993-01-27

    A series of proposed polyketide chain elongation intermediates has been synthesized as the N-acetylcysteamine (NAC) thioesters and incorporated into nargenicin (1) in [sup 13]C-labeled form by administration to both actively fermenting and resting cultures of Nocardia argentinesis. Thus, feeding of (2S,3R)-[2,3-[sup 13]C[sub 2

  15. Identification of a Polyketide Synthase Required for Alternariol (AOH) and Alternariol-9-Methyl Ether (AME) Formation in Alternaria alternata

    PubMed Central

    Saha, Debjani; Fetzner, Ramona; Burkhardt, Britta; Podlech, Joachim; Metzler, Manfred; Dang, Ha; Lawrence, Christopher; Fischer, Reinhard

    2012-01-01

    Alternaria alternata produces more than 60 secondary metabolites, among which alternariol (AOH) and alternariol-9-methyl ether (AME) are important mycotoxins. Whereas the toxicology of these two polyketide-based compounds has been studied, nothing is known about the genetics of their biosynthesis. One of the postulated core enzymes in the biosynthesis of AOH and AME is polyketide synthase (PKS). In a draft genome sequence of A. alternata we identified 10 putative PKS-encoding genes. The timing of the expression of two PKS genes, pksJ and pksH, correlated with the production of AOH and AME. The PksJ and PksH proteins are predicted to be 2222 and 2821 amino acids in length, respectively. They are both iterative type I reducing polyketide synthases. PksJ harbors a peroxisomal targeting sequence at the C-terminus, suggesting that the biosynthesis occurs at least partly in these organelles. In the vicinity of pksJ we found a transcriptional regulator, altR, involved in pksJ induction and a putative methyl transferase, possibly responsible for AME formation. Downregulation of pksJ and altR caused a large decrease of alternariol formation, suggesting that PksJ is the polyketide synthase required for the postulated Claisen condensations during the biosynthesis. No other enzymes appeared to be required. PksH downregulation affected pksJ expression and thus caused an indirect effect on AOH production. PMID:22792370

  16. Nonhost-specific phytotoxicity of the polyketide-derived toxin solanapyrone A produced by Ascochyta rabiei and Alternaria solani

    USDA-ARS?s Scientific Manuscript database

    Solanapyrone A is a polyketide-derived metabolite produced by Ascochyta rabiei and Alternaria solani, which are the most destructive necrotrophic pathogens of chickpea and potato/tomato, respectively. They belong to the Order Pleosporales within the Class Dothideomycetes, but are phylogenetically di...

  17. Local bulk physics from intersecting modular Hamiltonians

    NASA Astrophysics Data System (ADS)

    Kabat, Daniel; Lifschytz, Gilad

    2017-06-01

    We show that bulk quantities localized on a minimal surface homologous to a boundary region correspond in the CFT to operators that commute with the modular Hamiltonian associated with the boundary region. If two such minimal surfaces intersect at a point in the bulk then CFT operators which commute with both extended modular Hamiltonians must be localized at the intersection point. We use this to construct local bulk operators purely from CFT considerations, without knowing the bulk metric, using intersecting modular Hamiltonians. For conformal field theories at zero and finite temperature the appropriate modular Hamiltonians are known explicitly and we recover known expressions for local bulk observables.

  18. Modular passive solar heating system

    SciTech Connect

    Hunter, B.D.

    1985-03-19

    A modular passive solar energy storage system comprises a plurality of heat tubes which are arranged to form a flat plate solar collector and are releasably connected to a water reservoir by, and are part of, double-walled heat exchangers which penetrate to the water reservoir and enhance the heat transfer characteristics between the collector and the reservoir. The flat plate collector-heat exchanger disassembly, the collector housing, and the reservoir are integrated into a relatively light weight, unitary structural system in which the reservoir is a primary structural element. In addition to light weight, the system features high efficiency and ease of assembly and maintenance.

  19. Modular design attitude control system

    NASA Technical Reports Server (NTRS)

    Chichester, F. D.

    1984-01-01

    A sequence of single axismodels and a series of reduced state linear observers of minimum order are used to reconstruct inaccessible variables pertaining to the modular attitude control of a rigid body flexible suspension model of a flexible spacecraft. The single axis models consist of two, three, four, and five rigid bodies, each interconnected by a flexible shaft passing through the mass centers of the bodies. Modal damping is added to each model. Reduced state linear observers are developed for synthesizing the inaccessible modal state variables for each modal model.

  20. Modular design attitude control system

    NASA Technical Reports Server (NTRS)

    Chichester, F. D.

    1983-01-01

    Application of modular control techniques to the attitude control of a prototype flexible spacecraft and a prototype flexible space platform was further developed by determining numerical values for the physical parameters of a four body approximation of the MSFC/hybrid deployable truss incorporated in the space platform model, generating sensitivity coefficients for the model of the flexible spacecraft, evaluating the changes in the digital computer simulation of the flexible spacecraft resulting from the addition of another rigid body to the model and comparing attitude control effectiveness with actuators on more than one rigid body of the model with that for the case in which the actuators were restricted to one body.

  1. Cascading dynamics in modular networks

    NASA Astrophysics Data System (ADS)

    Galstyan, Aram; Cohen, Paul

    2007-03-01

    In this paper we study a simple cascading process in a structured heterogeneous population, namely, a network composed of two loosely coupled communities. We demonstrate that under certain conditions the cascading dynamics in such a network has a two-tiered structure that characterizes activity spreading at different rates in the communities. We study the dynamics of the model using both simulations and an analytical approach based on annealed approximation and obtain good agreement between the two. Our results suggest that network modularity might have implications in various applications, such as epidemiology and viral marketing.

  2. Integrated modular engine - Reliability assessment

    NASA Astrophysics Data System (ADS)

    Parsley, R. C.; Ward, T. B.

    1992-07-01

    A major driver in the increased interest in integrated modular engine configurations is the desire for ultra reliability for future rocket propulsion systems. The concept of configuring multiple sets of turbomachinery networked to multiple thrust chamber assemblies has been identified as an approach with potential to achieve significant reliability enhancement. This paper summarizes the results of a reliability study comparing networked systems vs. discrete engine installations, both with and without major module and engine redundancy. The study was conducted for gas generator, expander, and staged combustion cycles. The results are representative of either booster or upper-stage applications and are indicative of either plug or nonplug installation philosophies.

  3. Structural and functional analysis of two di-domain aromatase/cyclases from type II polyketide synthases

    PubMed Central

    Caldara-Festin, Grace; Jackson, David R.; Barajas, Jesus F.; Valentic, Timothy R.; Patel, Avinash B.; Aguilar, Stephanie; Nguyen, MyChi; Vo, Michael; Khanna, Avinash; Sasaki, Eita; Liu, Hung-wen; Tsai, Shiou-Chuan

    2015-01-01

    Aromatic polyketides make up a large class of natural products with diverse bioactivity. During biosynthesis, linear poly-β-ketone intermediates are regiospecifically cyclized, yielding molecules with defined cyclization patterns that are crucial for polyketide bioactivity. The aromatase/cyclases (ARO/CYCs) are responsible for regiospecific cyclization of bacterial polyketides. The two most common cyclization patterns are C7–C12 and C9–C14 cyclizations. We have previously characterized three monodomain ARO/CYCs: ZhuI, TcmN, and WhiE. The last remaining uncharacterized class of ARO/CYCs is the di-domain ARO/CYCs, which catalyze C7–C12 cyclization and/or aromatization. Di-domain ARO/CYCs can further be separated into two subclasses: “nonreducing” ARO/CYCs, which act on nonreduced poly-β-ketones, and “reducing” ARO/CYCs, which act on cyclized C9 reduced poly-β-ketones. For years, the functional role of each domain in cyclization and aromatization for di-domain ARO/CYCs has remained a mystery. Here we present what is to our knowledge the first structural and functional analysis, along with an in-depth comparison, of the nonreducing (StfQ) and reducing (BexL) di-domain ARO/CYCs. This work completes the structural and functional characterization of mono- and di-domain ARO/CYCs in bacterial type II polyketide synthases and lays the groundwork for engineered biosynthesis of new bioactive polyketides. PMID:26631750

  4. Revised Modularity Index to Measure Modularity of OSS Projects with Case Study of Freemind

    NASA Astrophysics Data System (ADS)

    WahjuRahardjoEmanuel, Andi; Jahja Surjawan, Daniel

    2012-12-01

    Open Source Software (OSS) Projects are gaining popularity worldwide. Studies by many researchers show that the important key success factor is modularity of the source code. This paper presents the revised Modularity Index which is a software metrics to measure the modularity level of a javabased OSS Projects. To show its effectiveness in analyzing OSS Project, the Modularity Index and its supporting software metrics are then used to analyze the evolution of Freemind mind mapping OSS Project. The analysis using Modularity Index and its supporting metrics shows the strength and weaknesses of the Freemind OSS Projects.

  5. The Iterative Structure Analysis of Montgomery Modular Multiplication

    NASA Astrophysics Data System (ADS)

    Jinbo, Wang

    2007-09-01

    Montgomery modular multiplication (MMM) plays a crucial role in the implementation of modular exponentiations of public-key cryptography. In this paper, we discuss the iterative structure and extend the iterative bound condition of MMM. It can be applied to complicated modular exponentiations. Based on the iterative condition of MMM, we can directly use non-modular additions, subtractions and even simple multiplications instead of the modular forms, which make modular exponentiation operation very efficient but more importantly iterative applicability of MMM.

  6. Decentralized and Modular Electrical Architecture

    NASA Astrophysics Data System (ADS)

    Elisabelar, Christian; Lebaratoux, Laurence

    2014-08-01

    This paper presents the studies made on the definition and design of a decentralized and modular electrical architecture that can be used for power distribution, active thermal control (ATC), standard inputs-outputs electrical interfaces.Traditionally implemented inside central unit like OBC or RTU, these interfaces can be dispatched in the satellite by using MicroRTU.CNES propose a similar approach of MicroRTU. The system is based on a bus called BRIO (Bus Réparti des IO), which is composed, by a power bus and a RS485 digital bus. BRIO architecture is made with several miniature terminals called BTCU (BRIO Terminal Control Unit) distributed in the spacecraft.The challenge was to design and develop the BTCU with very little volume, low consumption and low cost. The standard BTCU models are developed and qualified with a configuration dedicated to ATC, while the first flight model will fly on MICROSCOPE for PYRO actuations and analogue acquisitions. The design of the BTCU is made in order to be easily adaptable for all type of electric interface needs.Extension of this concept is envisaged for power conditioning and distribution unit, and a Modular PCDU based on BRIO concept is proposed.

  7. Compact stellarators with modular coils.

    PubMed

    Garabedian, P R

    2000-07-18

    Compact stellarator designs with modular coils and only two or three field periods are now available; these designs have both good stability and quasiaxial symmetry providing adequate transport for a magnetic fusion reactor. If the bootstrap current assumes theoretically predicted values a three field period configuration is optimal, but if that net current turns out to be lower, a device with two periods and just 12 modular coils might be better. There are also attractive designs with quasihelical symmetry and four or five periods whose properties depend less on the bootstrap current. Good performance requires that there be a satisfactory magnetic well in the vacuum field, which is a property lacking in a stellarator-tokamak hybrid that has been proposed for a proof of principle experiment. In this paper, we present an analysis of stability for these configurations that is based on a mountain pass theorem asserting that, if two solutions of the problem of magnetohydrodynamic equilibrium can be found, then there has to be an unstable solution. We compare results of our theory of equilibrium, stability, and transport with recently announced measurements from the large LHD experiment in Japan.

  8. Compact stellarators with modular coils

    PubMed Central

    Garabedian, P. R.

    2000-01-01

    Compact stellarator designs with modular coils and only two or three field periods are now available; these designs have both good stability and quasiaxial symmetry providing adequate transport for a magnetic fusion reactor. If the bootstrap current assumes theoretically predicted values a three field period configuration is optimal, but if that net current turns out to be lower, a device with two periods and just 12 modular coils might be better. There are also attractive designs with quasihelical symmetry and four or five periods whose properties depend less on the bootstrap current. Good performance requires that there be a satisfactory magnetic well in the vacuum field, which is a property lacking in a stellarator-tokamak hybrid that has been proposed for a proof of principle experiment. In this paper, we present an analysis of stability for these configurations that is based on a mountain pass theorem asserting that, if two solutions of the problem of magnetohydrodynamic equilibrium can be found, then there has to be an unstable solution. We compare results of our theory of equilibrium, stability, and transport with recently announced measurements from the large LHD experiment in Japan. PMID:10899993

  9. A modular BLSS simulation model

    NASA Technical Reports Server (NTRS)

    Rummel, John D.; Volk, Tyler

    1987-01-01

    A bioregenerative life support system (BLSS) for extraterrestrial use will be faced with coordination problems more acute than those in any ecosystem found on Earth. A related problem in BLSS design is providing an interface between the various life support processors, one that will allow for their coordination while still allowing for system expansion. A modular model is presented of a BLSS that interfaces system processors only with the material storage reservoirs, allowing those reservoirs to act as the principal buffers in the system and thus minimizing difficulties with processor coordination. The modular nature of the model allows independent development of the detailed submodels that exist within the model framework. Using this model, BLSS dynamics were investigated under normal conditions and under various failure modes. Partial and complete failures of various components, such as the waste processors or the plants themselves, drive transient responses in the model system, allowing the examination of the effectiveness of the system reservoirs as buffers. The results from simulations help to determine control strategies and BLSS design requirements. An evolved version could be used as an interactive control aid in a future BLSS.

  10. Learning modular policies for robotics.

    PubMed

    Neumann, Gerhard; Daniel, Christian; Paraschos, Alexandros; Kupcsik, Andras; Peters, Jan

    2014-01-01

    A promising idea for scaling robot learning to more complex tasks is to use elemental behaviors as building blocks to compose more complex behavior. Ideally, such building blocks are used in combination with a learning algorithm that is able to learn to select, adapt, sequence and co-activate the building blocks. While there has been a lot of work on approaches that support one of these requirements, no learning algorithm exists that unifies all these properties in one framework. In this paper we present our work on a unified approach for learning such a modular control architecture. We introduce new policy search algorithms that are based on information-theoretic principles and are able to learn to select, adapt and sequence the building blocks. Furthermore, we developed a new representation for the individual building block that supports co-activation and principled ways for adapting the movement. Finally, we summarize our experiments for learning modular control architectures in simulation and with real robots.

  11. Learning modular policies for robotics

    PubMed Central

    Neumann, Gerhard; Daniel, Christian; Paraschos, Alexandros; Kupcsik, Andras; Peters, Jan

    2014-01-01

    A promising idea for scaling robot learning to more complex tasks is to use elemental behaviors as building blocks to compose more complex behavior. Ideally, such building blocks are used in combination with a learning algorithm that is able to learn to select, adapt, sequence and co-activate the building blocks. While there has been a lot of work on approaches that support one of these requirements, no learning algorithm exists that unifies all these properties in one framework. In this paper we present our work on a unified approach for learning such a modular control architecture. We introduce new policy search algorithms that are based on information-theoretic principles and are able to learn to select, adapt and sequence the building blocks. Furthermore, we developed a new representation for the individual building block that supports co-activation and principled ways for adapting the movement. Finally, we summarize our experiments for learning modular control architectures in simulation and with real robots. PMID:24966830

  12. Modularity, noise, and natural selection.

    PubMed

    Marroig, Gabriel; Melo, Diogo A R; Garcia, Guilherme

    2012-05-01

    Most biological systems are formed by component parts that are to some degree interrelated. Groups of parts that are more associated among themselves and are relatively autonomous from others are called modules. One of the consequences of modularity is that biological systems usually present an unequal distribution of the genetic variation among traits. Estimating the covariance matrix that describes these systems is a difficult problem due to a number of factors such as poor sample sizes and measurement errors. We show that this problem will be exacerbated whenever matrix inversion is required, as in directional selection reconstruction analysis. We explore the consequences of varying degrees of modularity and signal-to-noise ratio on selection reconstruction. We then present and test the efficiency of available methods for controlling noise in matrix estimates. In our simulations, controlling matrices for noise vastly improves the reconstruction of selection gradients. We also perform an analysis of selection gradients reconstruction over a New World Monkeys skull database to illustrate the impact of noise on such analyses. Noise-controlled estimates render far more plausible interpretations that are in full agreement with previous results. © 2012 The Author(s). Evolution© 2012 The Society for the Study of Evolution.

  13. A modular approach toward extremely large apertures

    NASA Astrophysics Data System (ADS)

    Woods, A. A., Jr.

    1981-02-01

    Modular antenna construction can provide a significant increase in reflector aperture size over deployable reflectors. The modular approach allows reflective mesh surfaces to be supported by a minimum of structure. The kinematics of the selected deployable design approach were validated by the subscale demonstration model. Further design refinements on the module structural/joints and design optimization on intermodule joints are needed.

  14. Deployable modular mesh antenna - Concept and feasibility

    NASA Astrophysics Data System (ADS)

    Mitsugi, Jin; Yasaka, Tetsuo

    The feasibility of a 10m aperture deployable modular mesh antenna is evaluated by integrating the results of a statistical surface accuracy estimation and of surface shape adjustment experiments. It has been clarified that by combining seven 4m aperture modules, a 10m aperture deployable modular mesh antenna can be constructed, preserving the surface accuracy that is applicable to C band mission.

  15. Modular Building Institute 2000 Educational Showcase.

    ERIC Educational Resources Information Center

    Modular Building Inst., Charlottesville, VA.

    This publication contains brief articles concerned with modular school structures. The articles offer examples of such structures at actual schools. The articles in this issue are: (1) "Elementary K-8 Modular Courtyard"; (2) "School District #33, Chilliwack, BC"; (3) "New Elementary School for Briarwood, NY"; (4) "Addition to Queens Intermediate…

  16. Modular Buildings Are Here To Stay.

    ERIC Educational Resources Information Center

    Williams, Steven; Roman, Michael I.; Tiernan, Maury; Savage, Chuck; Airikka, Robert; Brosius, Jerry L.

    2000-01-01

    Presents several examples of modular building construction being used be school districts to support their need for more space, building flexibility, and enhancement of the learning environment. Comparisons with traditionally built school facilities are offered as are answers to commonly held myths concerning modular construction. (GR)

  17. Modular Construction: The Wave of the Future.

    ERIC Educational Resources Information Center

    Savage, Chuck

    1989-01-01

    Modular construction of school buildings offers speed of construction, with 100 percent contractor responsibility for the completed structures. Under negotiated terms, modular projects can be purchased outright or through long-term leasing arrangements that provide ownership at the end of the lease period. (MLF)

  18. Detectability thresholds of general modular graphs

    NASA Astrophysics Data System (ADS)

    Kawamoto, Tatsuro; Kabashima, Yoshiyuki

    2017-01-01

    We investigate the detectability thresholds of various modular structures in the stochastic block model. Our analysis reveals how the detectability threshold is related to the details of the modular pattern, including the hierarchy of the clusters. We show that certain planted structures are impossible to infer regardless of their fuzziness.

  19. 48 CFR 39.103 - Modular contracting.

    Code of Federal Regulations, 2013 CFR

    2013-10-01

    ... 48 Federal Acquisition Regulations System 1 2013-10-01 2013-10-01 false Modular contracting. 39.103 Section 39.103 Federal Acquisition Regulations System FEDERAL ACQUISITION REGULATION SPECIAL... extent practicable, use modular contracting to acquire major systems (see 2.101) of...

  20. 48 CFR 39.103 - Modular contracting.

    Code of Federal Regulations, 2011 CFR

    2011-10-01

    ... 48 Federal Acquisition Regulations System 1 2011-10-01 2011-10-01 false Modular contracting. 39.103 Section 39.103 Federal Acquisition Regulations System FEDERAL ACQUISITION REGULATION SPECIAL... extent practicable, use modular contracting to acquire major systems (see 2.101) of...

  1. A Modular Laser Graphics Projection System

    NASA Astrophysics Data System (ADS)

    Newswanger, Craig D.

    1984-05-01

    WED Enterprises has designed and built a modular projection system for the presentation of animated laser shows. This system was designed specifically for use in Disney theme shows. Its modular design allows it to be adapted to many show situations with simple hardware and software adjustments. The primary goals were superior animation, long life, low maintenance and stand alone operation.

  2. The relative efficiency of modular and non-modular networks of different size

    PubMed Central

    Tosh, Colin R.; McNally, Luke

    2015-01-01

    Most biological networks are modular but previous work with small model networks has indicated that modularity does not necessarily lead to increased functional efficiency. Most biological networks are large, however, and here we examine the relative functional efficiency of modular and non-modular neural networks at a range of sizes. We conduct a detailed analysis of efficiency in networks of two size classes: ‘small’ and ‘large’, and a less detailed analysis across a range of network sizes. The former analysis reveals that while the modular network is less efficient than one of the two non-modular networks considered when networks are small, it is usually equally or more efficient than both non-modular networks when networks are large. The latter analysis shows that in networks of small to intermediate size, modular networks are much more efficient that non-modular networks of the same (low) connective density. If connective density must be kept low to reduce energy needs for example, this could promote modularity. We have shown how relative functionality/performance scales with network size, but the precise nature of evolutionary relationship between network size and prevalence of modularity will depend on the costs of connectivity. PMID:25631996

  3. The Sponge-Derived Fijianolide Polyketide Class: Further Evaluation of Their Structural and Cytotoxicity Properties

    PubMed Central

    Johnson, Tyler A.; Tenney, Karen; Cichewicz, Robert H.; Morinaka, Brandon I.; White, Kimberly N.; Amagata, Taro; Subramanian, Balanehru; Media, Joseph; Mooberry, Susan L.; Valeriote, Frederick A.; Crews, Phillip

    2009-01-01

    The sponge derived polyketide macrolides fijianolides A (1) and B (2) (a.k.a. isolaulimalide and laulimalide) have taxol-like microtubule-stabilizing activity and the latter exhibits potent cytotoxicity. Insight on the biogeographical and phenotypic variations of Cacospongia mycofijiensis is presented that will enable future study of the biosynthetic pathway that produces the fijianolides. In addition to fijianolides A and B, six new fijianolides, D–I (7–12), were isolated, each with modifications to the C-20 side chain of the macrolide ring. Compounds 7–12 exhibited a range of in vitro activities against HCT-116 and MDA-MB-435 cell lines. Fijianolides 8 and 10 were shown to disrupt interphase and mitotic division but were less potent than 2. An in vivo evaluation of 2 using tumor-bearing SCID mice demonstrated significant inhibition of growth in HCT-116 tumors over 28 days. PMID:17622130

  4. Crystallization and preliminary crystallographic analysis of an octaketide-producing plant type III polyketide synthase

    SciTech Connect

    Morita, Hiroyuki; Kondo, Shin; Kato, Ryohei; Wanibuchi, Kiyofumi; Noguchi, Hiroshi; Sugio, Shigetoshi; Abe, Ikuro; Kohno, Toshiyuki

    2007-11-01

    Octaketide synthase from A. arborescens has been overexpressed in E. coli, purified and crystallized. Diffraction data have been collected to 2.6 Å. Octaketide synthase (OKS) from Aloe arborescens is a plant-specific type III polyketide synthase that produces SEK4 and SEK4b from eight molecules of malonyl-CoA. Recombinant OKS expressed in Escherichia coli was crystallized by the hanging-drop vapour-diffusion method. The crystals belonged to space group I422, with unit-cell parameters a = b = 110.2, c = 281.4 Å, α = β = γ = 90.0°. Diffraction data were collected to 2.6 Å resolution using synchrotron radiation at BL24XU of SPring-8.

  5. A two-step sulfation in antibiotic biosynthesis requires a type III polyketide synthase.

    PubMed

    Tang, Xiaoyu; Eitel, Kornelia; Kaysser, Leonard; Kulik, Andreas; Grond, Stephanie; Gust, Bertolt

    2013-10-01

    Caprazamycins (CPZs) belong to a group of liponucleoside antibiotics inhibiting the bacterial MraY translocase, an essential enzyme involved in peptidoglycan biosynthesis. We have recently identified analogs that are decorated with a sulfate group at the 2″-hydroxy of the aminoribosyl moiety, and we now report an unprecedented two-step sulfation mechanism during the biosynthesis of CPZs. A type III polyketide synthase (PKS) known as Cpz6 is used in the biosynthesis of a group of new triketide pyrones that are subsequently sulfated by an unusual 3'-phosphoadenosine-5'-phosphosulfate (PAPS)-dependent sulfotransferase (Cpz8) to yield phenolic sulfate esters, which serve as sulfate donors for a PAPS-independent arylsulfate sulfotransferase (Cpz4) to generate sulfated CPZs. This finding is to our knowledge the first demonstration of genuine sulfate donors for an arylsulfate sulfotransferase and the first report of a type III PKS to generate a chemical reagent in bacterial sulfate metabolism.

  6. Daldionin, an Unprecedented Binaphthyl Derivative, and Diverse Polyketide Congeners from a Fungal Orchid Endophyte.

    PubMed

    Barnes, Emma C; Jumpathong, Juangjun; Lumyong, Saisamorn; Voigt, Kerstin; Hertweck, Christian

    2016-03-18

    Thailand possesses a rich diversity of orchid species that, in turn, live in symbiosis with a wide variety of fungi. Such endophytes have the potential to produce secondary metabolites with bioactivity against orchid and/or human pathogens. The orchid-associated fungal strain Daldinia eschscholtzii was found to produce a diverse range of aromatic polyketides including the new naphthalene derivatives daldionin, nodulones B and C, and daldinones F and G along with eight known compounds. Daldionin possesses an unprecedented oxane-linked binaphthyl ring system. These compounds demonstrate the high diversity of structural variations that are constructed during fungal biosynthesis, and the results include important observations concerning the biosynthesis of binaphthyl derivatives. Daldionin was found to have weak antiproliferative activity against HUVEC and K-562 cell lines. All but one of the isolated compounds showed moderate antimicrobial activity towards at least one of the four tested microbial strains.

  7. Forazoline A: Marine-derived polyketide with antifungal in vivo efficacy

    SciTech Connect

    Wyche, Thomas P.; Piotrowski, Jeff S.; Hou, Yanpeng; Braun, Doug; Deshpande, Raamesh; McIlwain, Sean; Ong, Irene M.; Myers, Chad L.; Guzei, Ilia A.; Westler, William M.; Andes, David R.; Bugni, Tim S.

    2014-09-04

    Forazoline A, a novel antifungal polyketide with in vivo efficacy against Candida albicans, was discovered using LCMS-based metabolomics to investigate marine-invertebrate-associated bacteria. Forazoline A had a highly unusual and unprecedented skeleton. Acquisition of 13C–13C gCOSY and 13C–15N HMQC NMR data provided the direct carbon–carbon and carbon–nitrogen connectivity, respectively. This approach represents the first example of determining direct 13C–15N connectivity for a natural product. In this paper, using yeast chemical genomics, we propose that forazoline A operated through a new mechanism of action with a phenotypic outcome of disrupting membrane integrity.

  8. Six New Polyketide Decalin Compounds from Mangrove Endophytic Fungus Penicillium aurantiogriseum 328#

    PubMed Central

    Ma, Yanhong; Li, Jing; Huang, Meixiang; Liu, Lan; Wang, Jun; Lin, Yongcheng

    2015-01-01

    Six new compounds with polyketide decalin ring, peaurantiogriseols A–F (1–6), along with two known compounds, aspermytin A (7), 1-propanone,3-hydroxy-1-(1,2,4a,5,6,7,8,8a-octahydro-2,5-dihydroxy-1,2,6-trimethyl-1-naphthalenyl) (8), were isolated from the fermentation products of mangrove endophytic fungus Penicillium aurantiogriseum 328#. Their structures were elucidated based on their structure analysis. The absolute configurations of compounds 1 and 2 were determined by 1H NMR analysis of their Mosher esters; the absolute configurations of 3–6 were determined by using theoretical calculations of electronic circular dichroism (ECD). Compounds 1–8 showed low inhibitory activity against human aldose reductase, no activity of inducing neurite outgrowth, nor antimicrobial activity. PMID:26473887

  9. Engineering Fungal Nonreducing Polyketide Synthase by Heterologous Expression and Domain Swapping

    SciTech Connect

    Yeh, Hsu-Hua; Chang, Shu-Lin; Chiang, Yi-Ming; Bruno, Kenneth S.; Oakley, Berl R.; Wu, Tung-Kung; Wang, Clay C. C.

    2013-02-15

    Heterologous expression of the A. niger NR-PKS gene, e_gw1_19.204 and the adjacent stand-alone R domain gene, est_GWPlus_C_190476 in A. nidulans demonstrated that they belong to a single gene named dtbA. The DtbA protein produces two polyketides, 2,4-dihydroxy-3,5,6-trimethylbenzaldehyde 1 and 2-ethyl-4,6-dihydroxy-3,5-dimethylbenzaldehyde 2. Generation of DtbA+R-TE chimeric PKSs by swapping the DtbA R domain with the AusA (austinol biosynthesis) or ANID_06448 TE domain enabled the production of two metabolites with carboxylic acids replacing the corresponding aldehydes.

  10. An evolutionary model encompassing substrate specificity and reactivity of type I polyketide synthase thioesterases.

    PubMed

    Hari, Taylor P A; Labana, Puneet; Boileau, Meaghan; Boddy, Christopher N

    2014-12-15

    Bacterial polyketides are a rich source of chemical diversity and pharmaceutical agents. Understanding the biochemical basis for their biosynthesis and the evolutionary driving force leading to this diversity is essential to take advantage of the enzymes as biocatalysts and to access new chemical diversity for drug discovery. Biochemical characterization of the thioesterase (TE) responsible for 6-deoxyerythronolide macrocyclization shows that a small, evolutionarily accessible change to the substrate can increase the chemical diversity of products, including macrodiolide formation. We propose an evolutionary model in which TEs are by nature non-selective for the type of chemistry they catalyze, producing a range of metabolites. As one metabolite becomes essential for improving fitness in a particular environment, the TE evolves to enrich for that corresponding reactivity. This hypothesis is supported by our phylogenetic analysis, showing convergent evolution of macrodiolide-forming TEs.

  11. Polyketide and nonribosomal peptide retro-biosynthesis and global gene cluster matching.

    PubMed

    Dejong, Chris A; Chen, Gregory M; Li, Haoxin; Johnston, Chad W; Edwards, Mclean R; Rees, Philip N; Skinnider, Michael A; Webster, Andrew L H; Magarvey, Nathan A

    2016-12-01

    Polyketides (PKs) and nonribosomal peptides (NRPs) are profoundly important natural products, forming the foundations of many therapeutic regimes. Decades of research have revealed over 11,000 PK and NRP structures, and genome sequencing is uncovering new PK and NRP gene clusters at an unprecedented rate. However, only ∼10% of PK and NRPs are currently associated with gene clusters, and it is unclear how many of these orphan gene clusters encode previously isolated molecules. Therefore, to efficiently guide the discovery of new molecules, we must first systematically de-orphan emergent gene clusters from genomes. Here we provide to our knowledge the first comprehensive retro-biosynthetic program, generalized retro-biosynthetic assembly prediction engine (GRAPE), for PK and NRP families and introduce a computational pipeline, global alignment for natural products cheminformatics (GARLIC), to uncover how observed biosynthetic gene clusters relate to known molecules, leading to the identification of gene clusters that encode new molecules.

  12. Discovery of a novel superfamily of type III polyketide synthases in Aspergillus oryzae.

    PubMed

    Seshime, Yasuyo; Juvvadi, Praveen Rao; Fujii, Isao; Kitamoto, Katsuhiko

    2005-05-27

    Identification of genes encoding type III polyketide synthase (PKS) superfamily members in the industrially useful filamentous fungus, Aspergillus oryzae, revealed that their distribution is not specific to plants or bacteria. Among other Aspergilli (Aspergillus nidulans and Aspergillus fumigatus), A. oryzae was unique in possessing four chalcone synthase (CHS)-like genes (csyA, csyB, csyC, and csyD). Expression of csyA, csyB, and csyD genes was confirmed by RT-PCR. Comparative genome analyses revealed single putative type III PKS in Neurospora crassa and Fusarium graminearum, two each in Magnaporthe grisea and Podospora anserina, and three in Phenarocheate chrysosporium, with a phylogenic distinction from bacteria and plants. Conservation of catalytic residues in the CHSs across species implicated enzymatically active nature of these newly discovered homologs.

  13. Epothilone C macrolactonization and hydrolysis are catalyzed by the isolated thioesterase domain of epothilone polyketide synthase.

    PubMed

    Boddy, Christopher N; Schneider, Tanya L; Hotta, Kinya; Walsh, Christopher T; Khosla, Chaitan

    2003-03-26

    Epothilone C is produced by the combined action of one nonribosomal peptide synthetase (NRPS) and nine polyketide synthase (PKS) modules in a multienzyme system. The final step in the biosynthesis is the thioesterase (TE)-catalyzed cyclorelease of epothilone from the EpoF protein. It has been unclear whether isolated PKS TE domains could exhibit macrolactonization activity. Here we demonstrate that the excised epothilone TE domain can catalyze the efficient cyclization of the N-acetylcysteamine thioester of seco-epothilone C to generate epothilone C (kcat/KM = 0.41 +/- 0.03 min-1 mM-1). The TE domain also catalyzes the hydrolysis of both the N-acetylcysteamine thioester of seco-epothilone C (kcat = 0.087 +/- 0.005 min-1, KM = 291 +/- 53 muM) and that of the epothilone C (kcat = 0.67 +/- 0.01 min-1, KM = 117 +/- 5 muM) to form seco-epothilone C.

  14. Cytotoxic Polyketides from the Deep-Sea-Derived Fungus Engyodontium album DFFSCS021

    PubMed Central

    Yao, Qifeng; Wang, Jie; Zhang, Xiaoyong; Nong, Xuhua; Xu, Xinya; Qi, Shuhua

    2014-01-01

    Eight new chromones, engyodontiumones A–H (1–8), and three new phenol derivatives (9–11) together with eight known polyketides (12–19) were isolated from the deep-sea-derived fungus Engyodontium album DFFSCS021. Their structures were identified by extensive spectroscopic analysis. Compounds 8 and 16 showed significant selective cytotoxicity against human histiocytic lymphoma U937 cell line with IC50 values of 4.9 and 8.8 μM, respectively. In addition, this is the first time to report that 8, 15 and 16 had mild antibacterial activity against Escherichia coli and Bacillus subtilis, and 15 showed potent antilarval activity against barnacle Balanus amphitrite larval settlement. PMID:25501793

  15. Aspergiolides C and D: spirocyclic aromatic polyketides with potent protein kinase c-Met inhibitory effects.

    PubMed

    Du, Lin; Ai, Jing; Li, Dehai; Zhu, Tianjiao; Wang, Ying; Knauer, Michael; Bruhn, Torsten; Liu, Hongbing; Geng, Meiyu; Gu, Qianqun; Bringmann, Gerhard

    2011-01-24

    Variation of the cultivation conditions for Aspergillus glaucus led to the discovery of two novel spirocyclic aromatic polyketides, aspergiolides C (3) and D (4). Their constitutions were elucidated by a combination of spectroscopic methods and isotope-labeling experiments. Aspergiolides C (3) and D (4) occur as racemic mixtures, the resolution of which was succeeded by HPLC on a chiral phase. The absolute configurations of their enantiomers were assigned online, from the peaks in the chromatogram, by a combination of HPLC-CD and quantum chemical CD calculations. Both compounds were found to inhibit the kinase activities of the receptor tyrosine kinases (RTKs) c-Met, Ron, and c-Src with low-micromolar IC(50)s. The enantiomers of 3 were resolved by HPLC on a chiral phase. Both enantiomers showed a comparable inhibition of the HGF-induced autophosphorylation of c-Met and of subsequent cell migration.

  16. Modularity maximization using completely positive programming

    NASA Astrophysics Data System (ADS)

    Yazdanparast, Sakineh; Havens, Timothy C.

    2017-04-01

    Community detection is one of the most prominent problems of social network analysis. In this paper, a novel method for Modularity Maximization (MM) for community detection is presented which exploits the Alternating Direction Augmented Lagrangian (ADAL) method for maximizing a generalized form of Newman's modularity function. We first transform Newman's modularity function into a quadratic program and then use Completely Positive Programming (CPP) to map the quadratic program to a linear program, which provides the globally optimal maximum modularity partition. In order to solve the proposed CPP problem, a closed form solution using the ADAL merged with a rank minimization approach is proposed. The performance of the proposed method is evaluated on several real-world data sets used for benchmarks community detection. Simulation results shows the proposed technique provides outstanding results in terms of modularity value for crisp partitions.

  17. Finding network communities using modularity density

    NASA Astrophysics Data System (ADS)

    Botta, Federico; del Genio, Charo I.

    2016-12-01

    Many real-world complex networks exhibit a community structure, in which the modules correspond to actual functional units. Identifying these communities is a key challenge for scientists. A common approach is to search for the network partition that maximizes a quality function. Here, we present a detailed analysis of a recently proposed function, namely modularity density. We show that it does not incur in the drawbacks suffered by traditional modularity, and that it can identify networks without ground-truth community structure, deriving its analytical dependence on link density in generic random graphs. In addition, we show that modularity density allows an easy comparison between networks of different sizes, and we also present some limitations that methods based on modularity density may suffer from. Finally, we introduce an efficient, quadratic community detection algorithm based on modularity density maximization, validating its accuracy against theoretical predictions and on a set of benchmark networks.

  18. The emergence of modularity in biological systems

    NASA Astrophysics Data System (ADS)

    Lorenz, Dirk M.; Jeng, Alice; Deem, Michael W.

    2011-06-01

    In this review, we discuss modularity and hierarchy in biological systems. We review examples from protein structure, genetics, and biological networks of modular partitioning of the geometry of biological space. We review theories to explain modular organization of biology, with a focus on explaining how biology may spontaneously organize to a structured form. That is, we seek to explain how biology nucleated from among the many possibilities in chemistry. The emergence of modular organization of biological structure will be described as a symmetry-breaking phase transition, with modularity as the order parameter. Experimental support for this description will be reviewed. Examples will be presented from pathogen structure, metabolic networks, gene networks, and protein-protein interaction networks. Additional examples will be presented from ecological food networks, developmental pathways, physiology, and social networks.

  19. Functional Promiscuity of Two Divergent Paralogs of Type III Plant Polyketide Synthases1

    PubMed Central

    Pandith, Shahzad A.; Dhar, Niha; Bhat, Wajid Waheed; Kushwaha, Manoj; Gupta, Ajai P.; Shah, Manzoor A.; Vishwakarma, Ram

    2016-01-01

    Plants effectively defend themselves against biotic and abiotic stresses by synthesizing diverse secondary metabolites, including health-protective flavonoids. These display incredible chemical diversity and ubiquitous occurrence and confer impeccable biological and agricultural applications. Chalcone synthase (CHS), a type III plant polyketide synthase, is critical for flavonoid biosynthesis. It catalyzes acyl-coenzyme A thioesters to synthesize naringenin chalcone through a polyketidic intermediate. The functional divergence among the evolutionarily generated members of a gene family is pivotal in driving the chemical diversity. Against this backdrop, this study was aimed to functionally characterize members of the CHS gene family from Rheum emodi, an endangered and endemic high-altitude medicinal herb of northwestern Himalayas. Two full-length cDNAs (1,179 bp each), ReCHS1 and ReCHS2, encoding unique paralogs were isolated and characterized. Heterologous expression and purification in Escherichia coli, bottom-up proteomic characterization, high-performance liquid chromatography-electrospray ionization-tandem mass spectrometry analysis, and enzyme kinetic studies using five different substrates confirmed their catalytic potential. Phylogenetic analysis revealed the existence of higher synonymous mutations in the intronless divergents of ReCHS. ReCHS2 displayed significant enzymatic efficiency (Vmax/Km) with different substrates. There were significant spatial and altitudinal variations in messenger RNA transcript levels of ReCHSs correlating positively with metabolite accumulation. Furthermore, the elicitations in the form of methyl jasmonate, salicylic acid, ultraviolet B light, and wounding, chosen on the basis of identified cis-regulatory promoter elements, presented considerable differences in the transcript profiles of ReCHSs. Taken together, our results demonstrate differential propensities of CHS paralogs in terms of the accumulation of flavonoids and

  20. Reconstitution of the iterative type II polyketide synthase for tetracenomycin F2 biosynthesis.

    PubMed

    Bao, W; Wendt-Pienkowski, E; Hutchinson, C R

    1998-06-02

    The tetracenomycin polyketide synthase (TCM PKS), a type II complex, produces TCM F2, a precursor of TCM C in Streptomyces glaucescens, and consists of at least the TcmK, -L, -M, and -N proteins. The TcmK/TcmL ketosynthase subunits were purified from overexpression of their genes in Streptomyces lividans. TcmK (calculated molecular mass 45 kd) and TcmL (calculated molecular mass 42 kd) function as a heterodimeric alphabeta complex based on observing that only the purified complex complemented TCM PKS activity in protein extracts made from strains bearing tcmK or tcmL deletion mutants to make TCM F2 in vitro, and that the molecular mass of the purified complex was 90 kd as estimated by gel filtration chromatography. The TCM PKS activity was reconstituted with purified protein components, indicating that the minimal set of proteins required to make TCM F2 included the ketosynthase complex (TcmKL), an acyl carrier protein (TcmM), a malonyl CoA:ACP acyltransferase (MAT), and a cyclase (TcmN). The MAT was required to catalyze the transacylation between malonyl-CoA and TcmM, although a relatively slow spontaneous transacylation also occurred in a reaction without the MAT. Acetyl-CoA, the proposed starter unit for the TCM PKS, was not required for the production of TCM F2 in vitro, although it could be incorporated into this polyketide to a small extent. TcmJ, a PKS protein without a known function, greatly increased the production of TCM F2 but could not replace TcmN as a cyclase in the reconstituted system.

  1. Habitat-specific type I polyketide synthases in soils and street sediments.

    PubMed

    Hill, Patrick; Piel, Jörn; Aris-Brosou, Stéphane; Krištůfek, Václav; Boddy, Christopher N; Dijkhuizen, Lubbert

    2014-01-01

    Actinomycetes produce many pharmaceutically useful compounds through type I polyketide biosynthetic pathways. Soil has traditionally been an important source for these actinomycete-derived pharmaceuticals. As the rate of antibiotic discovery has decreased and the incidence of antibiotic resistance has increased, researchers have looked for alternatives to soil for bioprospecting. Street sediment, where actinomycetes make up a larger fraction of the bacterial population than in soil, is one such alternative environment. To determine if these differences in actinomycetal community structure are reflected in type I polyketide synthases (PKSI) distribution, environmental DNA from soils and street sediments was characterized by sequencing amplicons of PKSI-specific PCR primers. Amplicons covered two domains: the last 80 amino acids of the ketosynthase (KS) domain and the first 240 amino acids of the acyltransferase (AT) domain. One hundred and ninety clones from ten contrasting soils from six regions and nine street sediments from six cities were sequenced. Twenty-five clones from two earthworm-affected samples were also sequenced. UniFrac lineage-specific analysis identified two clades that clustered with actinomycetal GenBank matches that were street sediment-specific, one similar to the PKSI segment of the mycobactin siderophore involved in mycobacterial virulence. A clade of soil-specific sequences clustered with GenBank matches from the ambruticin and jerangolid pathways of Sorangium cellulosum. All three of these clades were found in sites >700 km apart. Street sediments are enriched in actinomycetal PKSIs. Non-actinomycetal PKSI pathways may be more chemically diverse than actinomycetal PKSIs. Common soil and street sediment PKIs are globally distributed.

  2. A family of polyketide synthase genes expressed in ripening Rubus fruits.

    PubMed

    Kumar, Amrita; Ellis, Brian E

    2003-02-01

    Quality traits of raspberry fruits such as aroma and color derive in part from the polyketide derivatives, benzalacetone and dihydrochalcone, respectively. The formation of these metabolites during fruit ripening is the result of the activity of polyketide synthases (PKS), benzalcetone synthase and chalcone synthase (CHS), during fruit development. To gain an understanding of the regulation of these multiple PKSs during fruit ripening, we have characterized the repertoire of Rubus PKS genes and studied their expression patterns during fruit ripening. Using a PCR-based homology search, a family of ten PKS genes (Ripks1-10) sharing 82-98% nucleotide sequence identity was identified in the Rubus idaeus genome. Low stringency screening of a ripening fruit-specific cDNA library, identified three groups of PKS cDNAs. Group 1 and 2 cDNAs were also represented in the PCR amplified products, while group 3 represented a new class of Rubus PKS gene. The Rubus PKS gene-family thus consists of at least eleven members. The three cDNAs exhibit distinct tissue-specific and developmentally regulated patterns of expression. RiPKS5 has high constitutive levels of expression in all organs, including developing flowers and fruits, while RiPKS6 and RiPKS11 expression is consistent with developmental and tissue-specific regulation in various organs. The recombinant proteins encoded by the three RiPKS cDNAs showed a typical CHS-type PKS activity. While phylogenetic analysis placed the three Rubus PKSs in one cluster, suggesting a recent duplication event, their distinct expression patterns suggest that their regulation, and thus function(s), has evolved independently of the structural genes themselves.

  3. Identification and functional characterization of three type III polyketide synthases from Aquilaria sinensis calli.

    PubMed

    Wang, Xiaohui; Zhang, Zhongxiu; Dong, Xianjuan; Feng, Yingying; Liu, Xiao; Gao, Bowen; Wang, Jinling; Zhang, Le; Wang, Juan; Shi, Shepo; Tu, Pengfei

    2017-03-30

    Type III polyketide synthases (PKSs) play an important role in biosynthesis of various plant secondary metabolites and plant adaptation to environmental stresses. Aquilaria sinensis is the main plant species for production of agarwood, little is known about the PKS family. In this study, AsCHS1 and two new type III PKSs, AsPKS1 and AsPKS2, were isolated and characterized in Aquilaria sinensis calli. The comparative sequence and phylogenetic analysis indicated that AsPKS1 and AsPKS2 belong to non-CHS group different from AsCHS1. The recombinant AsPKS1 and AsPKS2 produced the lactone-type products, suggesting different enzyme activities with AsCHS1. Three PKS genes had a tissues-specific pattern in A. sinensis. Moreover, we examined the expression profiles of three PKS genes in calli under different abiotic stresses and hormone treatments. AsCHS1 transcript was significantly induced by salt stress, AsPKS1 abundance was most remarkably enhanced by CdCl2 treatment, while AsPKS2 expression was most significantly induced by mannitol treatment. Furthermore, AsCHS1, AsPKS1 and AsPKS2 transcript were enhanced upon gibberellins (GA3), methyl jasmonate (MeJA), salicylic acid (SA) treatments, while three PKS genes displayed low transcript levels at the early stage under abscisic acid (ABA) treatment. In addition, three GFP:PKSs fusion proteins were localized in the cytoplasm and cell wall in Nicotiana benthamiana cells. These results indicated the multifunctional role of three type III PKSs in polyketide biosynthesis, plant resistance in abiotic stresses and signal transduction.

  4. Intimate bacterial-fungal interaction triggers biosynthesis of archetypal polyketides in Aspergillus nidulans.

    PubMed

    Schroeckh, Volker; Scherlach, Kirstin; Nützmann, Hans-Wilhelm; Shelest, Ekaterina; Schmidt-Heck, Wolfgang; Schuemann, Julia; Martin, Karin; Hertweck, Christian; Brakhage, Axel A

    2009-08-25

    Fungi produce numerous low molecular weight molecules endowed with a multitude of biological activities. However, mining the full-genome sequences of fungi indicates that their potential to produce secondary metabolites is greatly underestimated. Because most of the biosynthesis gene clusters are silent under laboratory conditions, one of the major challenges is to understand the physiological conditions under which these genes are activated. Thus, we cocultivated the important model fungus Aspergillus nidulans with a collection of 58 soil-dwelling actinomycetes. By microarray analyses of both Aspergillus secondary metabolism and full-genome arrays and Northern blot and quantitative RT-PCR analyses, we demonstrate at the molecular level that a distinct fungal-bacterial interaction leads to the specific activation of fungal secondary metabolism genes. Most surprisingly, dialysis experiments and electron microscopy indicated that an intimate physical interaction of the bacterial and fungal mycelia is required to elicit the specific response. Gene knockout experiments provided evidence that one induced gene cluster codes for the long-sought after polyketide synthase (PKS) required for the biosynthesis of the archetypal polyketide orsellinic acid, the typical lichen metabolite lecanoric acid, and the cathepsin K inhibitors F-9775A and F-9775B. A phylogenetic analysis demonstrates that orthologs of this PKS are widespread in nature in all major fungal groups, including mycobionts of lichens. These results provide evidence of specific interaction among microorganisms belonging to different domains and support the hypothesis that not only diffusible signals but intimate physical interactions contribute to the communication among microorganisms and induction of otherwise silent biosynthesis genes.

  5. Probing the Selectivity and Protein•Protein Interactions of a Non-Reducing Fungal Polyketide Synthase Using Mechanism-Based Crosslinkers

    PubMed Central

    Bruegger, Joel; Haushalter, Bob; Vagstad, Anna; Shakya, Gaurav; Mih, Nathan; Townsend, Craig A.; Burkart, Michael D.; Tsai, Shiou-Chuan

    2013-01-01

    SUMMARY Protein•protein interactions, which often involve interactions between an acyl carrier protein (ACP) and its partner enzymes, are important for coordinating polyketide biosynthesis. However, the nature of such interactions is not well understood, especially in the fungal non-reducing polyketide synthases (NR-PKSs) that biosynthesize toxic and pharmaceutically important polyketides. Here, we employ a mechanism-based crosslinker to successfully probe ACP and ketosynthase (KS) domain interactions in NR-PKSs. We found that crosslinking efficiency is closely correlated with the strength of ACP•KS interactions, and that KS demonstrates strong starter unit selectivity. We further identified positively charged surface residues by KS mutagenesis, which mediate key interactions with the negatively-charged ACP surface. Such complementary/matching contact pairs can serve as “adapter surfaces” for future efforts to generate new polyketides using NR-PKSs. PMID:23993461

  6. Biosynthesis of Akaeolide and Lorneic Acids and Annotation of Type I Polyketide Synthase Gene Clusters in the Genome of Streptomyces sp. NPS554

    PubMed Central

    Zhou, Tao; Komaki, Hisayuki; Ichikawa, Natsuko; Hosoyama, Akira; Sato, Seizo; Igarashi, Yasuhiro

    2015-01-01

    The incorporation pattern of biosynthetic precursors into two structurally unique polyketides, akaeolide and lorneic acid A, was elucidated by feeding experiments with 13C-labeled precursors. In addition, the draft genome sequence of the producer, Streptomyces sp. NPS554, was performed and the biosynthetic gene clusters for these polyketides were identified. The putative gene clusters contain all the polyketide synthase (PKS) domains necessary for assembly of the carbon skeletons. Combined with the 13C-labeling results, gene function prediction enabled us to propose biosynthetic pathways involving unusual carbon-carbon bond formation reactions. Genome analysis also indicated the presence of at least ten orphan type I PKS gene clusters that might be responsible for the production of new polyketides. PMID:25603349

  7. Small Modular Reactors: Institutional Assessment

    SciTech Connect

    Joseph Perkowski, Ph.D.

    2012-06-01

    ? Objectives include, among others, a description of the basic development status of “small modular reactors” (SMRs) focused primarily on domestic activity; investigation of the domestic market appeal of modular reactors from the viewpoints of both key energy sector customers and also key stakeholders in the financial community; and consideration of how to proceed further with a pro-active "core group" of stakeholders substantially interested in modular nuclear deployment in order to provide the basis to expedite design/construction activity and regulatory approval. ? Information gathering was via available resources, both published and personal communications with key individual stakeholders; published information is limited to that already in public domain (no confidentiality); viewpoints from interviews are incorporated within. Discussions at both government-hosted and private-hosted SMR meetings are reflected herein. INL itself maintains a neutral view on all issues described. Note: as per prior discussion between INL and CAP, individual and highly knowledgeable senior-level stakeholders provided the bulk of insights herein, and the results of those interviews are the main source of the observations of this report. ? Attachment A is the list of individual stakeholders consulted to date, including some who provided significant earlier assessments of SMR institutional feasibility. ? Attachments B, C, and D are included to provide substantial context on the international status of SMR development; they are not intended to be comprehensive and are individualized due to the separate nature of the source materials. Attachment E is a summary of the DOE requirements for winning teams regarding the current SMR solicitation. Attachment F deserves separate consideration due to the relative maturity of the SMART SMR program underway in Korea. Attachment G provides illustrative SMR design features and is intended for background. Attachment H is included for overview

  8. A bioinspired modular aquatic robot

    NASA Astrophysics Data System (ADS)

    Tallapragada, Phanindra; Pollard, Beau

    2016-11-01

    Several bio inspired swimming robots exist which seek to emulate the morphology of fish and the flapping motion of the tail and fins or other appendages and body of aquatic creatures. The locomotion of such robots and the aquatic animals that they seek to emulate is determined to a large degree by the changes in the shape of the body, which produce periodic changes in the momentum of the body and the creation and interaction of the vorticity field in the fluid with the body. We demonstrate an underactuated robot which swims due to the periodic changes in the angular momentum of the robot effected by the motion of an internal rotor. The robot is modular, unactuated tail like segments can be easily added to the robot. These segments modulate the interaction of the body with the fluid to produce a variety of passive shape changes that can allow the robot to swim in different modes.

  9. Analytical Spectroscopy Using Modular Systems

    NASA Astrophysics Data System (ADS)

    Patterson, Brian M.; Danielson, Neil D.; Lorigan, Gary A.; Sommer, André J.

    2003-12-01

    This article describes the development of three analytical spectroscopy experiments that compare the determination of salicylic acid (SA) content in aspirin tablets. The experiments are based on UV vis, fluorescence, and Raman spectroscopies and utilize modular spectroscopic components. Students assemble their own instruments, optimize them with respect to signal-to-noise, generate calibration curves, determine the SA content in retail aspirin tablets, and assign features in the respective spectra to functional groups within the active material. Using this approach in the discovery-based setting, the students gain invaluable insight into method-specific parameters, such as instrumental components, sample preparation, and analytical capability. In addition, the students learn the fundamentals of fiber optics and signal processing using the low-cost CCD based spectroscopic components.

  10. BESST: A Miniature, Modular Radiometer

    NASA Technical Reports Server (NTRS)

    Warden, Robert; Good, William; Baldwin-Stevens, Erik

    2010-01-01

    A new radiometer assembly has been developed that incorporates modular design principles in order to provide flexibility and versatility. The assembly, shown in Figure 1, is made up of six modules plus a central cubical frame. A small thermal imaging detector is used to determine the temperature of remote objects. To improve the accuracy of the temperature reading, frequent calibration is required. The detector must view known temperature targets before viewing the remote object. Calibration is achieved by using a motorized fold mirror to select the desired scene the detector views. The motor steps the fold mirror through several positions, which allows the detector to view the calibration targets or the remote object. The details, features, and benefits of the radiometer are described in this paper.

  11. MODULAR MANIPULATOR FOR ROBOTICS APPLICATIONS

    SciTech Connect

    Joseph W. Geisinger, Ph.D.

    2001-07-31

    ARM Automation, Inc. is developing a framework of modular actuators that can address the DOE's wide range of robotics needs. The objective of this effort is to demonstrate the effectiveness of this technology by constructing a manipulator from these actuators within a glovebox for Automated Plutonium Processing (APP). At the end of the project, the system of actuators was used to construct several different manipulator configurations, which accommodate common glovebox tasks such as repackaging. The modular nature and quickconnects of this system simplify installation into ''hot'' boxes and any potential modifications or repair therein. This work focused on the development of self-contained robotic actuator modules including the embedded electronic controls for the purpose of building a manipulator system. Both of the actuators developed under this project contain the control electronics, sensors, motor, gear train, wiring, system communications and mechanical interfaces of a complete robotics servo device. Test actuators and accompanying DISC{trademark}s underwent validation testing at The University of Texas at Austin and ARM Automation, Inc. following final design and fabrication. The system also included custom links, an umbilical cord, an open architecture PC-based system controller, and operational software that permitted integration into a completely functional robotic manipulator system. The open architecture on which this system is based avoids proprietary interfaces and communication protocols which only serve to limit the capabilities and flexibility of automation equipment. The system was integrated and tested in the contractor's facility for intended performance and operations. The manipulator was tested using the full-scale equipment and process mock-ups. The project produced a practical and operational system including a quantitative evaluation of its performance and cost.

  12. Osmotrophy in modular Ediacara organisms

    PubMed Central

    Laflamme, Marc; Xiao, Shuhai; Kowalewski, Michał

    2009-01-01

    The Ediacara biota include macroscopic, morphologically complex soft-bodied organisms that appear globally in the late Ediacaran Period (575–542 Ma). The physiology, feeding strategies, and functional morphology of the modular Ediacara organisms (rangeomorphs and erniettomorphs) remain debated but are critical for understanding their ecology and phylogeny. Their modular construction triggered numerous hypotheses concerning their likely feeding strategies, ranging from micro-to-macrophagus feeding to photoautotrophy to osmotrophy. Macrophagus feeding in rangeomorphs and erniettomorphs is inconsistent with their lack of oral openings, and photoautotrophy in rangeomorphs is contradicted by their habitats below the photic zone. Here, we combine theoretical models and empirical data to evaluate the feasibility of osmotrophy, which requires high surface area to volume (SA/V) ratios, as a primary feeding strategy of rangeomorphs and erniettomorphs. Although exclusively osmotrophic feeding in modern ecosystems is restricted to microscopic bacteria, this study suggests that (i) fractal branching of rangeomorph modules resulted in SA/V ratios comparable to those observed in modern osmotrophic bacteria, and (ii) rangeomorphs, and particularly erniettomorphs, could have achieved osmotrophic SA/V ratios similar to bacteria, provided their bodies included metabolically inert material. Thus, specific morphological adaptations observed in rangeomorphs and erniettomorphs may have represented strategies for overcoming physiological constraints that typically make osmotrophy prohibitive for macroscopic life forms. These results support the viability of osmotrophic feeding in rangeomorphs and erniettomorphs, help explain their taphonomic peculiarities, and point to the possible importance of earliest macroorganisms for cycling dissolved organic carbon that may have been present in abundance during Ediacaran times. PMID:19706530

  13. Size reduction of complex networks preserving modularity

    NASA Astrophysics Data System (ADS)

    Arenas, A.; Duch, J.; Fernández, A.; Gómez, S.

    2007-06-01

    The ubiquity of modular structure in real-world complex networks is the focus of attention in many trials to understand the interplay between network topology and functionality. The best approaches to the identification of modular structure are based on the optimization of a quality function known as modularity. However this optimization is a hard task provided that the computational complexity of the problem is in the non-deterministic polynomial-time hard (NP-hard) class. Here we propose an exact method for reducing the size of weighted (directed and undirected) complex networks while maintaining their modularity. This size reduction allows use of heuristic algorithms that optimize modularity for a better exploration of the modularity landscape. We compare the modularity obtained in several real complex-networks by using the extremal optimization algorithm, before and after the size reduction, showing the improvement obtained. We speculate that the proposed analytical size reduction could be extended to an exact coarse graining of the network in the scope of real-space renormalization.

  14. A Prototype for Modular Cell Engineering.

    PubMed

    Wilbanks, Brandon; Layton, Donovan; Garcia, Sergio; Trinh, Cong

    2017-10-10

    When aiming to produce a target chemical at high yield, titer, and productivity, various combinations of genetic parts available to build the target pathway can generate a large number of strains for characterization. This engineering approach will become increasingly laborious and expensive when seeking to develop desirable strains for optimal production of a large space of biochemicals due to extensive screening. Our recent theoretical development of modular cell (MODCELL) design principles can offer a promising solution for rapid generation of optimal strains by coupling a modular cell and exchangeable production modules in a plug-and-play fashion. In this study, we experimentally validated some designed properties of MODCELL by demonstrating: i) a modular (chassis) cell is required to couple with a production module, a heterologous ethanol pathway, as a testbed, ii) degree of coupling between the modular cell and production modules can be modulated to enhance growth and product synthesis, iii) a modular cell can be used as a host to select an optimal pyruvate decarboxylase (PDC) of the ethanol production module and to help identify a hypothetical PDC protein, and iv) adaptive laboratory evolution based on growth selection of the modular cell can enhance growth and product synthesis rates. We envision that the MODCELL design provides a powerful prototype for modular cell engineering to rapidly create optimal strains for synthesis of a large space of biochemicals.

  15. Size reduction of complex networks preserving modularity

    SciTech Connect

    Arenas, A.; Duch, J.; Fernandez, A.; Gomez, S.

    2008-12-24

    The ubiquity of modular structure in real-world complex networks is being the focus of attention in many trials to understand the interplay between network topology and functionality. The best approaches to the identification of modular structure are based on the optimization of a quality function known as modularity. However this optimization is a hard task provided that the computational complexity of the problem is in the NP-hard class. Here we propose an exact method for reducing the size of weighted (directed and undirected) complex networks while maintaining invariant its modularity. This size reduction allows the heuristic algorithms that optimize modularity for a better exploration of the modularity landscape. We compare the modularity obtained in several real complex-networks by using the Extremal Optimization algorithm, before and after the size reduction, showing the improvement obtained. We speculate that the proposed analytical size reduction could be extended to an exact coarse graining of the network in the scope of real-space renormalization.

  16. Rational design of efficient modular cells.

    PubMed

    Trinh, Cong T; Liu, Yan; Conner, David J

    2015-11-01

    The modular cell design principle is formulated to devise modular (chassis) cells. These cells can be assembled with exchangeable production modules in a plug-and-play fashion to build microbial cell factories for efficient combinatorial biosynthesis of novel molecules, requiring minimal iterative strain optimization steps. A modular cell is designed to be auxotrophic, containing core metabolic pathways that are necessary but insufficient to support cell growth and maintenance. To be functional, it must tightly couple with an exchangeable production module containing auxiliary metabolic pathways that not only complement cell growth but also enhance production of targeted molecules. We developed a MODCELL (modular cell) framework based on metabolic pathway analysis to implement the modular cell design principle. MODCELL identifies genetic modifications and requirements to construct modular cell candidates and their associated exchangeable production modules. By defining the degree of similarity and coupling metrics, MODCELL can evaluate which exchangeable production module(s) can be tightly coupled with a modular cell candidate. We first demonstrated how MODCELL works in a step-by-step manner for example metabolic networks, and then applied it to design modular Escherichia coli cells for efficient combinatorial biosynthesis of five alcohols (ethanol, propanol, isopropanol, butanol and isobutanol) and five butyrate esters (ethyl butyrate, propyl butyrate, isopropyl butyrate, butyl butyrate and isobutyl butyrate) from pentose sugars (arabinose and xylose) and hexose sugars (glucose, mannose, and galactose) under anaerobic conditions. We identified three modular cells, MODCELL1, MODCELL2 and MODCELL3, that can couple well with Group 1 of modules (ethanol, isobutanol, butanol, ethyl butyrate, isobutyl butyrate, butyl butyrate), Group 2 (isopropanol, isopropyl butyrate), and Group 3 (propanol, isopropanol), respectively. We validated the design of MODCELL1 for anaerobic

  17. Generalized epidemic process on modular networks

    NASA Astrophysics Data System (ADS)

    Chung, Kihong; Baek, Yongjoo; Kim, Daniel; Ha, Meesoon; Jeong, Hawoong

    2014-05-01

    Social reinforcement and modular structure are two salient features observed in the spreading of behavior through social contacts. In order to investigate the interplay between these two features, we study the generalized epidemic process on modular networks with equal-sized finite communities and adjustable modularity. Using the analytical approach originally applied to clique-based random networks, we show that the system exhibits a bond-percolation type continuous phase transition for weak social reinforcement, whereas a discontinuous phase transition occurs for sufficiently strong social reinforcement. Our findings are numerically verified using the finite-size scaling analysis and the crossings of the bimodality coefficient.

  18. A Modular Approach to Redundant Robot Control

    SciTech Connect

    Anderson, R.J.

    1997-12-01

    This paper describes a modular approach for computing redundant robot kinematics. First some conventional redundant control methods are presented and shown to be `passive control laws`, i.e. they can be represented by a network consisting of passive elements. These networks are then put into modular form by applying scattering operator techniques. Additional subnetwork modules can then be added to further shape the motion. Modules for obstacle detection, joint limit avoidance, proximity sensing, and for imposing nonlinear velocity constraints are presented. The resulting redundant robot control system is modular, flexible and robust.

  19. The gravity duals of modular Hamiltonians

    NASA Astrophysics Data System (ADS)

    Jafferis, Daniel L.; Suh, S. Josephine

    2016-09-01

    In this work, we investigate modular Hamiltonians defined with respect to arbitrary spatial regions in quantum field theory states which have semi-classical gravity duals. We find prescriptions in the gravity dual for calculating the action of the modular Hamiltonian on its defining state, including its dual metric, and also on small excitations around the state. Curiously, use of the covariant holographic entanglement entropy formula leads us to the conclusion that the modular Hamiltonian, which in the quantum field theory acts only in the causal completion of the region, does not commute with bulk operators whose entire gauge-invariant description is space-like to the causal completion of the region.

  20. The gravity duals of modular Hamiltonians

    SciTech Connect

    Jafferis, Daniel L.; Suh, S. Josephine

    2016-09-12

    In this study, we investigate modular Hamiltonians defined with respect to arbitrary spatial regions in quantum field theory states which have semi-classical gravity duals. We find prescriptions in the gravity dual for calculating the action of the modular Hamiltonian on its defining state, including its dual metric, and also on small excitations around the state. Curiously, use of the covariant holographic entanglement entropy formula leads us to the conclusion that the modular Hamiltonian, which in the quantum field theory acts only in the causal completion of the region, does not commute with bulk operators whose entire gauge-invariant description is space-like to the causal completion of the region.

  1. Deciphering Piperidine Formation in Polyketide-Derived Indolizidines Reveals a Thioester Reduction, Transamination, and Unusual Imine Reduction Process.

    PubMed

    Peng, Haidong; Wei, Erman; Wang, Jiali; Zhang, Yanan; Cheng, Lin; Ma, Hongmin; Deng, Zixin; Qu, Xudong

    2016-12-16

    Piperidine and indolizidine are two basic units of alkaloids that are frequently observed in natural and synthetic compounds. Their biosynthesis in natural products is highly conserved and mostly derived from the incorporation of lysine cyclization products. Through in vitro reconstitution, we herein identified a novel pathway involving a group of polyketide-derived indolizidines, which comprises the processes of tandem two-electron thioester reduction, transamination, and imine reduction to convert acyl carrier protein (ACP)-tethered polyketide chains into the piperidine moieties of their indolizidine scaffolds. The enzymes that catalyze the imine reduction are distinct from previous known imine reductases, which have a fold of acyl-CoA dehydrogenase but do not require flavin for reduction. Our results not only provide a new way for the biosynthesis of the basic units of alkaloids but also show a novel class of imine reductases that may benefit the fields of biocatalysis and biomanufacturing.

  2. Divergent Mechanistic Routes for the Formation of gem-Dimethyl Groups in the Biosynthesis of Complex Polyketides

    SciTech Connect

    Poust, S; Phelan, RM; Deng, K; Katz, L; Petzold, CJ; Keasling, JD

    2015-01-07

    The gem-dimethyl groups in polyketide-derived natural products add steric bulk and, accordingly, lend increased stability to medicinal compounds, however, our ability to rationally incorporate this functional group in modified natural products is limited. In order to characterize the mechanism of gem-dimethyl group formation, with a goal toward engineering of novel compounds containing this moiety, the gem-dimethyl group producing polyketide synthase (PKS) modules of yersiniabactin and epothilone were characterized using mass spectrometry. The work demonstrated, contrary to the canonical understanding of reaction order in PKSs, that methylation can precede condensation in gem-dimethyl group producing PKS modules. Experiments showed that both PKSs are able to use dimethylmalonyl acyl carrier protein (ACP) as an extender unit. Interestingly, for epothilone module8, use of dimethylmalonyl-ACP appeared to be the sole route to form a gem-dimethylated product, while the yersiniabactin PKS could methylate before or after ketosynthase condensation.

  3. A novel erythromycin, 6-desmethyl erythromycin D, made by substituting an acyltransferase domain of the erythromycin polyketide synthase.

    PubMed

    Petkovic, Hrvoje; Lill, Rachel E; Sheridan, Rose M; Wilkinson, Barrie; McCormick, Ellen L; McArthur, Hamish A I; Staunton, James; Leadlay, Peter F; Kendrew, Steven G

    2003-06-01

    The acyltransferase (AT) domain in module 4 of the erythromycin polyketide synthase (PKS) was substituted with an AT domain from the rapamycin PKS module 2 in order to alter the substrate specificity from methylmalonyl-CoA to malonyl-CoA. The resulting strain produced 6-desmethyl erythromycin D as the predominant product. This AT domain swap completes the library of malonyl-CoA AT swaps on the erythromycin PKS and reinforces PKS engineering as a robust and generic tool.

  4. Synthesis of isoxazoles en route to semi-aromatized polyketides: dehydrogenation of benzonitrile oxide-para-quinone acetal cycloadducts.

    PubMed

    Hashimoto, Yoshimitsu; Takada, Akiomi; Takikawa, Hiroshi; Suzuki, Keisuke

    2012-08-14

    A variety of highly functionalized polycyclic isoxazoles are prepared by a two-step protocol: (1) 1,3-dipolar cycloaddition of o,o'-disubstituted benzonitrile oxides to para-quinone mono-acetals, then (2) dehydrogenation. The cycloaddition proceeds in a regioselective manner, favouring the formation of the 4-acyl cycloadducts, which are suitable intermediates for the synthesis of semi-aromatized polycyclic targets derived from polyketide type-II biosynthesis.

  5. Modular optimization code package: MOZAIK

    NASA Astrophysics Data System (ADS)

    Bekar, Kursat B.

    This dissertation addresses the development of a modular optimization code package, MOZAIK, for geometric shape optimization problems in nuclear engineering applications. MOZAIK's first mission, determining the optimal shape of the D2O moderator tank for the current and new beam tube configurations for the Penn State Breazeale Reactor's (PSBR) beam port facility, is used to demonstrate its capabilities and test its performance. MOZAIK was designed as a modular optimization sequence including three primary independent modules: the initializer, the physics and the optimizer, each having a specific task. By using fixed interface blocks among the modules, the code attains its two most important characteristics: generic form and modularity. The benefit of this modular structure is that the contents of the modules can be switched depending on the requirements of accuracy, computational efficiency, or compatibility with the other modules. Oak Ridge National Laboratory's discrete ordinates transport code TORT was selected as the transport solver in the physics module of MOZAIK, and two different optimizers, Min-max and Genetic Algorithms (GA), were implemented in the optimizer module of the code package. A distributed memory parallelism was also applied to MOZAIK via MPI (Message Passing Interface) to execute the physics module concurrently on a number of processors for various states in the same search. Moreover, dynamic scheduling was enabled to enhance load balance among the processors while running MOZAIK's physics module thus improving the parallel speedup and efficiency. In this way, the total computation time consumed by the physics module is reduced by a factor close to M, where M is the number of processors. This capability also encourages the use of MOZAIK for shape optimization problems in nuclear applications because many traditional codes related to radiation transport do not have parallel execution capability. A set of computational models based on the

  6. Crystallization and preliminary X-ray diffraction studies of polyketide synthase-1 (PKS-1) from Cannabis sativa

    SciTech Connect

    Taguchi, Chiho; Taura, Futoshi; Tamada, Taro; Shoyama, Yoshinari; Shoyama, Yukihiro; Tanaka, Hiroyuki; Kuroki, Ryota; Morimoto, Satoshi

    2008-03-01

    Polyketide synthase-1 from C. sativa has been crystallized. The crystal diffracted to 1.55 Å resolution with sufficient quality for further structure determination. Polyketide synthase-1 (PKS-1) is a novel type III polyketide synthase that catalyzes the biosynthesis of hexanoyl triacetic acid lactone in Cannabis sativa (Mexican strain). PKS-1 was overproduced in Escherichia coli, purified and finally crystallized in two different space groups. The crystal obtained in 0.1 M HEPES buffer pH 7.5 containing 0.2 M calcium acetate and 20%(w/v) polyethylene glycol 3350 diffracted to 1.65 Å resolution and belonged to space group P1, with unit-cell parameters a = 54.3, b = 59.3, c = 62.6 Å, α = 69, β = 81, γ = 80°. Another crystal obtained in 0.1 M HEPES buffer pH 7.5 containing 0.2 M sodium chloride and 20%(w/v) polyethylene glycol 3350 diffracted to 1.55 Å resolution and belonged to space group P2{sub 1}2{sub 1}2{sub 1}, with unit-cell parameters a = 54.3, b = 110, c = 130 Å. These data will enable us to determine the crystal structure of PKS-1.

  7. Functional Gene-Guided Discovery of Type II Polyketides from Culturable Actinomycetes Associated with Soft Coral Scleronephthya sp

    PubMed Central

    Sun, Wei; Peng, Chongsheng; Zhao, Yunyu; Li, Zhiyong

    2012-01-01

    Compared with the actinomycetes in stone corals, the phylogenetic diversity of soft coral-associated culturable actinomycetes is essentially unexplored. Meanwhile, the knowledge of the natural products from coral-associated actinomycetes is very limited. In this study, thirty-two strains were isolated from the tissue of the soft coral Scleronephthya sp. in the East China Sea, which were grouped into eight genera by 16S rDNA phylogenetic analysis: Micromonospora, Gordonia, Mycobacterium, Nocardioides, Streptomyces, Cellulomonas, Dietzia and Rhodococcus. 6 Micromonospora strains and 4 Streptomyces strains were found to be with the potential for producing aromatic polyketides based on the analysis of KSα (ketoacyl-synthase) gene in the PKS II (type II polyketides synthase) gene cluster. Among the 6 Micromonospora strains, angucycline cyclase gene was amplified in 2 strains (A5-1 and A6-2), suggesting their potential in synthesizing angucyclines e.g. jadomycin. Under the guidance of functional gene prediction, one jadomycin B analogue (7b, 13-dihydro-7-O-methyl jadomycin B) was detected in the fermentation broth of Micromonospora sp. strain A5-1. This study highlights the phylogenetically diverse culturable actinomycetes associated with the tissue of soft coral Scleronephthya sp. and the potential of coral-derived actinomycetes especially Micromonospora in producing aromatic polyketides. PMID:22880121

  8. Systematic Domain Swaps of Iterative, Nonreducing Polyketide Synthases Provide a Mechanistic Understanding and Rationale For Catalytic Reprogramming

    PubMed Central

    2015-01-01

    Iterative, nonreducing polyketide synthases (NR-PKSs) are multidomain enzymes responsible for the construction of the core architecture of aromatic polyketide natural products in fungi. Engineering these enzymes for the production of non-native metabolites has been a long-standing goal. We conducted a systematic survey of in vitro “domain swapped” NR-PKSs using an enzyme deconstruction approach. The NR-PKSs were dissected into mono- to multidomain fragments and recombined as noncognate pairs in vitro, reconstituting enzymatic activity. The enzymes used in this study produce aromatic polyketides that are representative of the four main chemical features set by the individual NR-PKS: starter unit selection, chain-length control, cyclization register control, and product release mechanism. We found that boundary conditions limit successful chemistry, which are dependent on a set of underlying enzymatic mechanisms. Crucial for successful redirection of catalysis, the rate of productive chemistry must outpace the rate of spontaneous derailment and thioesterase-mediated editing. Additionally, all of the domains in a noncognate system must interact efficiently if chemical redirection is to proceed. These observations refine and further substantiate current understanding of the mechanisms governing NR-PKS catalysis. PMID:24815013

  9. Identification of a Polyketide Synthase Gene in the Synthesis of Phleichrome of the Phytopathogenic Fungus Cladosporium phlei

    PubMed Central

    So, Kum-Kang; Chung, Yun-Jo; Kim, Jung-Mi; Kim, Beom-Tae; Park, Seung-Moon; Kim, Dae-Hyuk

    2015-01-01

    Phleichrome, a pigment produced by the phytopathogenic fungus Cladosporium phlei, is a fungal perylenequinone whose photodynamic activity has been studied intensively. To determine the biological function of phleichrome and to engineer a strain with enhanced production of phleichrome, we identified the gene responsible for the synthesis of phleichrome. Structural comparison of phleichrome with other fungal perylenequinones suggested that phleichrome is synthesized via polyketide pathway. We recently identified four different polyketide synthase (PKS) genes encompassing three major clades of fungal PKSs that differ with respect to reducing conditions for the polyketide product. Based on in silico analysis of cloned genes, we hypothesized that the non-reducing PKS gene, Cppks1, is involved in phleichrome biosynthesis. Increased accumulation of Cppks1 transcript was observed in response to supplementation with the application of synthetic inducer cyclo-(l-Pro-l-Phe). In addition, heterologous expression of the Cppks1 gene in Cryphonectria parasitica resulted in the production of phleichrome. These results provide convincing evidence that the Cppks1 gene is responsible for the biosynthesis of phleichrome. PMID:26612679

  10. The CTB1 gene encoding a fungal polyketide synthase is required for cercosporin biosynthesis and fungal virulence of Cercospora nicotianae.

    PubMed

    Choquer, Mathias; Dekkers, Katherine L; Chen, Hui-Qin; Cao, Lihua; Ueng, Peter P; Daub, Margaret E; Chung, Kuang-Ren

    2005-05-01

    Cercosporin is a light-activated, non-host-selective toxin produced by many Cercospora fungal species. In this study, a polyketide synthase gene (CTB1) was functionally identified and molecularly characterized to play a key role in cercosporin biosynthesis by Cercospora nicotianae. We also provide conclusive evidence to confirm the crucial role of cercosporin in fungal pathogenesis. CTB1 encoded a polypeptide with a deduced length of 2,196 amino acids containing a keto synthase (KS), an acyltransferase (AT), a thioesterase/claisen cyclase (TE/CYC), and two acyl carrier protein (ACP) domains, and had high levels of similarity to many fungal type I polyketide synthases. Expression of a 6.8-kb CTB1 transcript was highly regulated by light and medium composition, consistent with the conditions required for cercosporin biosynthesis in cultures. Targeted disruption of CTB1 resulted in the loss of both CTB1 transcript and cercosporin biosynthesis in C. nicotianae. The ctb1-null mutants incited fewer necrotic lesions on inoculated tobacco leaves compared with the wild type. Complementation of ctb1-null mutants with a full-length CTB1 clone restored wild-type levels of cercosporin production as well as the ability to induce lesions on tobacco. Thus, we have demonstrated conclusively that cercosporin is synthesized via a polyketide pathway, and cercosporin is an important virulence factor in C. nicotianae. The results also suggest that strategies that avoid the toxicity of cercosporin will be useful in reduction of disease incidence caused by Cercospora spp.

  11. Crystal structure and biochemical studies of the trans-acting polyketide enoyl reductase LovC from lovastatin biosynthesis

    PubMed Central

    Ames, Brian D.; Nguyen, Chi; Bruegger, Joel; Smith, Peter; Xu, Wei; Ma, Suzanne; Wong, Emily; Wong, Steven; Xie, Xinkai; Li, Jesse W.-H.; Vederas, John C.; Tang, Yi; Tsai, Shiou-Chuan

    2012-01-01

    Lovastatin is an important statin prescribed for the treatment and prevention of cardiovascular diseases. Biosynthesis of lovastatin uses an iterative type I polyketide synthase (PKS). LovC is a trans-acting enoyl reductase (ER) that specifically reduces three out of eight possible polyketide intermediates during lovastatin biosynthesis. Such trans-acting ERs have been reported across a variety of other fungal PKS enzymes as a strategy in nature to diversify polyketides. How LovC achieves such specificity is unknown. The 1.9-Å structure of LovC reveals that LovC possesses a medium-chain dehydrogenase/reductase (MDR) fold with a unique monomeric assembly. Two LovC cocrystal structures and enzymological studies help elucidate the molecular basis of LovC specificity, define stereochemistry, and identify active-site residues. Sequence alignment indicates a general applicability to trans-acting ERs of fungal PKSs, as well as their potential application to directing biosynthesis. PMID:22733743

  12. Anatomy of the β-branching enzyme of polyketide biosynthesis and its interaction with an acyl-ACP substrate

    PubMed Central

    Maloney, Finn P.; Gerwick, Lena; Gerwick, William H.; Sherman, David H.; Smith, Janet L.

    2016-01-01

    Alkyl branching at the β position of a polyketide intermediate is an important variation on canonical polyketide natural product biosynthesis. The branching enzyme, 3-hydroxy-3-methylglutaryl synthase (HMGS), catalyzes the aldol addition of an acyl donor to a β-keto-polyketide intermediate acceptor. HMGS is highly selective for two specialized acyl carrier proteins (ACPs) that deliver the donor and acceptor substrates. The HMGS from the curacin A biosynthetic pathway (CurD) was examined to establish the basis for ACP selectivity. The donor ACP (CurB) had high affinity for the enzyme (Kd = 0.5 μM) and could not be substituted by the acceptor ACP. High-resolution crystal structures of HMGS alone and in complex with its donor ACP reveal a tight interaction that depends on exquisite surface shape and charge complementarity between the proteins. Selectivity is explained by HMGS binding to an unusual surface cleft on the donor ACP, in a manner that would exclude the acceptor ACP. Within the active site, HMGS discriminates between pre- and postreaction states of the donor ACP. The free phosphopantetheine (Ppant) cofactor of ACP occupies a conserved pocket that excludes the acetyl-Ppant substrate. In comparison with HMG-CoA (CoA) synthase, the homologous enzyme from primary metabolism, HMGS has several differences at the active site entrance, including a flexible-loop insertion, which may account for the specificity of one enzyme for substrates delivered by ACP and the other by CoA. PMID:27573844

  13. Policing starter unit selection of the enterocin type II polyketide synthase by the type II thioesterase EncL.

    PubMed

    Kalaitzis, John A; Cheng, Qian; Meluzzi, Dario; Xiang, Longkuan; Izumikawa, Miho; Dorrestein, Pieter C; Moore, Bradley S

    2011-11-15

    Enterocin is an atypical type II polyketide synthase (PKS) product from the marine actinomycete 'Streptomyces maritimus'. The enterocin biosynthesis gene cluster (enc) codes for proteins involved in the assembly and attachment of the rare benzoate primer that initiates polyketide assembly with the addition of seven malonate molecules and culminates in a Favorskii-like rearrangement of the linear poly-β-ketone to give its distinctive non-aromatic, caged core structure. Fundamental to enterocin biosynthesis, which utilizes a single acyl carrier protein (ACP), EncC, for both priming with benzoate and elongating with malonate, involves maintaining the correct balance of acyl-EncC substrates for efficient polyketide assembly. Here, we report the characterization of EncL as a type II thioesterase that functions to edit starter unit (mis)priming of EncC. We performed a series of in vivo mutational studies, heterologous expression experiments, in vitro reconstitution studies, and Fourier-transform mass spectrometry-monitored competitive enzyme assays that together support the proposed selective hydrolase activity of EncL toward misprimed acetyl-ACP over benzoyl-ACP to facilitate benzoyl priming of the enterocin PKS complex. While this system resembles the R1128 PKS that also utilizes an editing thioesterase (ZhuC) to purge acetate molecules from its initiation module ACP in favor of alkylacyl groups, the enterocin system is distinct in its usage of a single ACP for both priming and elongating reactions with different substrates. Copyright © 2011 Elsevier Ltd. All rights reserved.

  14. Insect-specific polyketide synthases (PKSs), potential PKS-nonribosomal peptide synthetase hybrids, and novel PKS clades in tropical fungi.

    PubMed

    Amnuaykanjanasin, Alongkorn; Phonghanpot, Suranat; Sengpanich, Nattapong; Cheevadhanarak, Supapon; Tanticharoen, Morakot

    2009-06-01

    Polyketides draw much attention because of their potential use in pharmaceutical and biotechnological applications. This study identifies an abundant pool of polyketide synthase (PKS) genes from local isolates of tropical fungi found in Thailand in three different ecological niches: insect pathogens, marine inhabitants, and lichen mutualists. We detected 149 PKS genes from 48 fungi using PCR with PKS-specific degenerate primers. We identified and classified 283 additional PKS genes from 13 fungal genomes. Phylogenetic analysis of all these PKS sequences the comprising ketosynthase (KS) conserved region and the KS-acyltransferase interdomain region yielded results very similar to those for phylogenies of the KS domain and suggested a number of remarkable points. (i) Twelve PKS genes amplified from 12 different insect-pathogenic fungi form a tight cluster, although along with two PKS genes extracted from genomes of Aspergillus niger and Aspergillus terreus, in reducing clade III. Some of these insect-specific fungal PKSs are nearly identical. (ii) We identified 38 new PKS-nonribosomal peptide synthetase hybrid genes in reducing clade II. (iii) Four distinct clades were discovered with more than 75% bootstrap support. We propose to designate the novel clade D1 with 100% bootstrap support "reducing clade V." The newly cloned PKS genes from these tropical fungi should provide useful and diverse genetic resources for future research on the characterization of polyketide compounds synthesized by these enzymes.

  15. Polyketide Synthase Gene Diversity within the Microbiome of the Sponge Arenosclera brasiliensis, Endemic to the Southern Atlantic Ocean

    PubMed Central

    Trindade-Silva, Amaro E.; Rua, Cintia P. J.; Andrade, Bruno G. N.; Vicente, Ana Carolina Paulo; Silva, Genivaldo G. Z.

    2013-01-01

    Microbes associated with marine sponges are considered important producers of bioactive, structurally unique polyketides. The synthesis of such secondary metabolites involves type I polyketide synthases (PKSs), which are enzymes that reach a maximum complexity degree in bacteria. The Haplosclerida sponge Arenosclera brasiliensis hosts a complex microbiota and is the source of arenosclerins, alkaloids with cytotoxic and antibacterial activity. In the present investigation, we performed high-throughput sequencing of the ketosynthase (KS) amplicon to investigate the diversity of PKS genes present in the metagenome of A. brasiliensis. Almost 4,000 ketosynthase reads were recovered, with about 90% annotated automatically as bacterial. A total of 235 bacterial KS contigs was rigorously assembled from this sequence pool and submitted to phylogenetic analysis. A great diversity of six type I PKS groups has been consistently detected in our phylogenetic reconstructions, including a novel and A. brasiliensis-exclusive group. Our study is the first to reveal the diversity of type I PKS genes in A. brasiliensis as well as the potential of its microbiome to serve as a source of new polyketides. PMID:23275501

  16. The polyketide synthase gene pks4 is essential for sexual development and regulates fruiting body morphology in Sordaria macrospora.

    PubMed

    Schindler, Daniel; Nowrousian, Minou

    2014-07-01

    Filamentous ascomycetes have long been known as producers of a variety of secondary metabolites, many of which have toxic effects on other organisms. However, the role of these metabolites in the biology of the fungi that produce them remains in most cases enigmatic. A major group of fungal secondary metabolites are polyketides. They are chemically diverse, but have in common that their chemical scaffolds are synthesized by polyketide synthases (PKSs). In a previous study, we analyzed development-dependent expression of pks genes in the filamentous ascomycete Sordaria macrospora. Here, we show that a deletion mutant of the pks4 gene is sterile, producing only protoperithecia but no mature perithecia, whereas overexpression of pks4 leads to enlarged, malformed fruiting bodies. Thus, correct expression levels of pks4 are essential for wild type-like perithecia formation. The predicted PKS4 protein has a domain structure that is similar to homologs in other fungi, but conserved residues of a methyl transferase domain present in other fungi are mutated in PKS4. Expression of several developmental genes is misregulated in the pks4 mutant. Surprisingly, the development-associated app gene is not downregulated in the mutant, in contrast to all other previously studied mutants with a block at the protoperithecial stage. Our data show that the polyketide synthase gene pks4 is essential for sexual development and plays a role in regulating fruiting body morphology. Copyright © 2014 Elsevier Inc. All rights reserved.

  17. Analysis of the Sequences, Structures, and Functions of Product-Releasing Enzyme Domains in Fungal Polyketide Synthases.

    PubMed

    Liu, Lu; Zhang, Zheng; Shao, Chang-Lun; Wang, Chang-Yun

    2017-01-01

    Product-releasing enzyme (PRE) domains in fungal non-reducing polyketide synthases (NR-PKSs) play a crucial role in catalysis and editing during polyketide biosynthesis, especially accelerating final biosynthetic reactions accompanied with product offloading. However, up to date, the systematic knowledge about PRE domains is deficient. In the present study, the relationships between sequences, structures, and functions of PRE domains were analyzed with 574 NR-PKSs of eight groups (I-VIII). It was found that the PRE domains in NR-PKSs could be mainly classified into three types, thioesterase (TE), reductase (R), and metallo-β-lactamase-type TE (MβL-TE). The widely distributed TE or TE-like domains were involved in NR-PKSs of groups I-IV, VI, and VIII. The R domains appeared in NR-PKSs of groups IV and VII, while the physically discrete MβL-TE domains were employed by most NR-PKSs of group V. The changes of catalytic sites and structural characteristics resulted in PRE functional differentiations. The phylogeny revealed that the evolution of TE domains was accompanied by complex functional divergence. The diverse sequence lengths of TE lid-loops affected substrate specificity with different chain lengths. The volume diversification of TE catalytic pockets contributed to catalytic mechanisms with functional differentiations. The above findings may help to understand the crucial catalysis of fungal aromatic polyketide biosyntheses and govern recombination of NR-PKSs to obtain unnatural target products.

  18. A putative polyketide-synthesis protein XC5357 from Xanthomonas campestris: heterologous expression, crystallization and preliminary X-ray analysis

    SciTech Connect

    Chu, Chiao-Li; Chin, Ko-Hsin; Lin, Fu-Yang; Chou, Chia-Cheng; Lee, Cheng-Chung; Shr, Hui-Lin; Lyu, Ping-Chiang; Wang, Andrew H.-J.; Chou, Shan-Ho

    2005-07-01

    A putative polyketide-synthesis protein XC5357 from X. campestris pv. campestris has been overexpressed in E. coli, purified and crystallized. The crystals diffracted to a resolution of at least 1.85 Å. Xanthomonas campestris pv. campestris (Xcc) is a Gram-negative yellow-pigmented bacterium and is the causative agent of black rot, one of the major worldwide diseases of cruciferous crops. It also synthesizes a variety of polyketide metabolites that lead to important antibiotics. XC5357 is a putative 12.2 kDa protein of unknown structure from Xcc that is likely to be essential for polyketide synthesis. It was overexpressed in Escherichia coli, purified and crystallized using the hanging-drop vapour-diffusion method. The crystals belong to the triclinic space group P1, with unit-cell parameters a = 43.7, b = 43.7, c = 46.5 Å, α = 65.0, β = 64.9, γ = 73.4°, and diffracted to a resolution of 1.85 Å.

  19. Modular Heat Exchanger With Integral Heat Pipe

    NASA Technical Reports Server (NTRS)

    Schreiber, Jeffrey G.

    1992-01-01

    Modular heat exchanger with integral heat pipe transports heat from source to Stirling engine. Alternative to heat exchangers depending on integrities of thousands of brazed joints, contains only 40 brazed tubes.

  20. Modular Solar Electric Power (MSEP) Systems (Presentation)

    SciTech Connect

    Hassani, V.

    2000-06-18

    This presentation discusses the development and deployment of Modular Solar Electric Power (MSEP) systems, the feasibility of application of existing binary power cycles to solar trough technology, and identification of next action items.

  1. Modular digital holographic fringe data processing system

    NASA Technical Reports Server (NTRS)

    Downward, J. G.; Vavra, P. C.; Schebor, F. S.; Vest, C. M.

    1985-01-01

    A software architecture suitable for reducing holographic fringe data into useful engineering data is developed and tested. The results, along with a detailed description of the proposed architecture for a Modular Digital Fringe Analysis System, are presented.

  2. The Modular Structure of Protein Networks

    NASA Astrophysics Data System (ADS)

    Rozenfeld, Hernán D.; Rybski, Diego; Havlin, Shlomo; Makse, Hernán A.

    2008-03-01

    The evolution of the human protein homology network (H-PHN) has led to a complex network that exhibits a surprisingly high level of modularity. Topologically, the H-PHN presents well connected groups (conformed by proteins of similar aminoacid structure) and weak connectivities between the groups. Here, we perform an empirical study of the H-PHN to characterize the degree of modularity in terms of scale-invariant laws using recently introduced box covering algorithms. We find that the exponent that determines the scale-invariance of the modularity is unexpectedly higher than the box dimension of the network. In addition, we perform a percolation analysis that gives insight into the evolutionary process that led to the modular organization and dynamics of the present H-PHN.

  3. Modular solar-heating system - design package

    NASA Technical Reports Server (NTRS)

    Sinton, D. S.

    1979-01-01

    Compilation contains design, performance, and hardware specifications in sufficient detail to fabricate or procure materials and install, operate, and maintain complete modular solar heating and hot water system for single family size dwellings.

  4. 47 CFR 15.212 - Modular transmitters.

    Code of Federal Regulations, 2012 CFR

    2012-10-01

    ... modular transmitter must have their own shielding. The physical crystal and tuning capacitors may be... shielded. The physical crystal and tuning capacitors may be located external to the shielded radio elements...

  5. Modular biowaste monitoring system conceptual design

    NASA Technical Reports Server (NTRS)

    Fogal, G. L.

    1974-01-01

    The objective of the study was to define requirements and generate a conceptual design for a Modular Biowaste Monitoring System for specifically supporting shuttle life science experimental and diagnostic programs.

  6. Modular, Intelligent Power Systems for Space Exploration

    NASA Technical Reports Server (NTRS)

    Button, Robert

    2006-01-01

    NASA's new Space Exploration Initiative demands that vehicles, habitats, and rovers achieve unprecedented levels of reliability, safety, effectiveness, and affordability. Modular and intelligent electrical power systems are critical to achieving those goals. Modular electrical power systems naturally increase reliability and safety through built-in fault tolerance. These modular systems also enable standardization across a multitude of systems, thereby greatly increasing affordability of the programs. Various technologies being developed to support this new paradigm for space power systems will be presented. Examples include the use of digital control in power electronics to enable better performance and advanced modularity functions such as distributed, master-less control and series input power conversion. Also, digital control and robust communication enables new levels of power system control, stability, fault detection, and health management. Summary results from recent development efforts are presented along with expected future technology development needs required to support NASA's ambitious space exploration goals.

  7. Modular modelling with Physiome standards.

    PubMed

    Cooling, Michael T; Nickerson, David P; Nielsen, Poul M F; Hunter, Peter J

    2016-12-01

    The complexity of computational models is increasing, supported by research in modelling tools and frameworks. But relatively little thought has gone into design principles for complex models. We propose a set of design principles for complex model construction with the Physiome standard modelling protocol CellML. By following the principles, models are generated that are extensible and are themselves suitable for reuse in larger models of increasing complexity. We illustrate these principles with examples including an architectural prototype linking, for the first time, electrophysiology, thermodynamically compliant metabolism, signal transduction, gene regulation and synthetic biology. The design principles complement other Physiome research projects, facilitating the application of virtual experiment protocols and model analysis techniques to assist the modelling community in creating libraries of composable, characterised and simulatable quantitative descriptions of physiology. The ability to produce and customise complex computational models has great potential to have a positive impact on human health. As the field develops towards whole-cell models and linking such models in multi-scale frameworks to encompass tissue, organ, or organism levels, reuse of previous modelling efforts will become increasingly necessary. Any modelling group wishing to reuse existing computational models as modules for their own work faces many challenges in the context of construction, storage, retrieval, documentation and analysis of such modules. Physiome standards, frameworks and tools seek to address several of these challenges, especially for models expressed in the modular protocol CellML. Aside from providing a general ability to produce modules, there has been relatively little research work on architectural principles of CellML models that will enable reuse at larger scales. To complement and support the existing tools and frameworks, we develop a set of principles to

  8. Optimal Network Modularity for Information Diffusion

    NASA Astrophysics Data System (ADS)

    Nematzadeh, Azadeh; Ferrara, Emilio; Flammini, Alessandro; Ahn, Yong-Yeol

    2014-08-01

    We investigate the impact of community structure on information diffusion with the linear threshold model. Our results demonstrate that modular structure may have counterintuitive effects on information diffusion when social reinforcement is present. We show that strong communities can facilitate global diffusion by enhancing local, intracommunity spreading. Using both analytic approaches and numerical simulations, we demonstrate the existence of an optimal network modularity, where global diffusion requires the minimal number of early adopters.

  9. Modular Programming Techniques for Distributed Computing Tasks

    DTIC Science & Technology

    2004-08-01

    Modular Programming Techniques for Distributed Computing Tasks Anthony Cowley, Hwa-Chow Hsu, Camillo J. Taylor GRASP Laboratory University of...network, distributed computing , software design 1. INTRODUCTION As efforts to field sensor networks, or teams of mobile robots, become more...TITLE AND SUBTITLE Modular Programming Techniques for Distributed Computing Tasks 5a. CONTRACT NUMBER 5b. GRANT NUMBER 5c. PROGRAM ELEMENT NUMBER

  10. Adaptive Coupled Oscillators for Modular Robots

    NASA Astrophysics Data System (ADS)

    Hartono, Pitoyo; Nakane, Aito

    In this research we physically built several robotics modules that are able to self-discover a connection topology which allows them to generate a coordinated behavior as an integrated modular robot. We consider that this self-configurability of hardware module can potentially simplify the costly designing process of complicated robots and at the same time improve the resiliency of modular robots in the face of internal and external changes.

  11. [Importance of neurobiology for modular psychotherapy].

    PubMed

    Schmahl, C; Bohus, M

    2013-11-01

    In the context of continuing education in psychiatry and psychotherapy, modular psychotherapy is of special importance. In modular psychotherapy, general interventions, e.g. for regulation of emotions, have an important function. In this review examples are given to describe the importance of neurobiology for the understanding and the improvement of these mechanisms. In addition, the use of neurobiological investigations within classical psychotherapy trials in the fields of borderline personality disorder and posttraumatic stress disorder will be depicted.

  12. A 3-d modular gripper design tool

    SciTech Connect

    Brown, R.G.; Brost, R.C.

    1997-01-01

    Modular fixturing kits are precisely machined sets of components used for flexible, short-turnaround construction of fixtures for a variety of manufacturing purposes. A modular vise is a parallel-jaw vise, where each jaw is a modular fixture plate with a regular grid of precisely positioned holes. A modular vise can be used to locate and hold parts for machining, assembly, and inspection tasks. To fixture a part, one places pins in some of the holes so that when the vise is closed, the part is reliably located and completely constrained. The modular vise concept can be adapted easily to the design of modular parallel-jaw grippers for robots. By attaching a grid plate to each jaw of a parallel-jaw gripper, the authors gain the ability to easily construct high-quality grasps for a wide variety of parts from a standard set of hardware. Wallack and Canny developed a previous algorithm for planning planar grasp configurations for the modular vise. In this paper, the authors expand this work to produce a 3-d fixture/gripper design tool. They describe several analyses added to the planar algorithm to improve its utility, including a three-dimensional grasp quality metric based on geometric and force information, three-dimensional geometric loading analysis, and inter-gripper interference analysis to determine the compatibility of multiple grasps for handing the part from one gripper to another. Finally, the authors describe two applications which combine the utility of modular vise-style grasping with inter-gripper interference: The first is the design of a flexible part-handling subsystem for a part cleaning workcell under development at Sandia National Laboratories; the second is the automatic design of grippers that support the assembly of multiple products on a single assembly line.

  13. Identification and characterization of the polyketide synthase involved in ochratoxin A biosynthesis in Aspergillus carbonarius

    SciTech Connect

    Gallo, Antonia; Knox, Benjamin P.; Bruno, Kenneth S.; Solfrizzo, Michele; Baker, Scott E.; Perrone, Giancarlo

    2014-06-02

    Ochratoxin A (OTA) is a potent mycotoxin produced by Aspergillus and Penicillium species and is a common contaminant of a wide variety of food commodities, with Aspergillus carbonarius being the main producer of OTA contamination in grapes and wine. The molecular structure of OTA is composed of a dihydroisocoumarin ring linked to phenylalanine and, as shown in different producing fungal species, a polyketide synthase (PKS) is a component of the OTA biosynthetic pathway. Similar to observations in other filamentous ascomycetes, the genome sequence of A. carbonarius contains a large number of genes predicted to encode PKSs. In this work a pks gene identified within the putative OTA cluster of A. carbonarius, designated as AcOTApks, was inactivated and the resulting mutant strain was unable to produce OTA, confirming the role of AcOTApks in this biosynthetic pathway. AcOTApks protein is characteristic of the highly reduced (HR)-PKS family, and also contains a putative methyltransferase domain likely responsible for the addition of the methyl group to the OTA polyketide structure. AcOTApks is different from the ACpks protein that we previously described which showed an expression profile compatible with OTA production. We performed phylogenetic analyses of the β-ketosynthase and acyl-transferase domains of the OTA PKSs which had been identified and characterized in different OTA producing fungal species. The phylogenetic results were similar for both the two domains analyzed and showed that OTA PKS of A. carbonarius, Aspergillus niger, and Aspergillus ochraceus clustered in a monophyletic group with 100% bootstrap support suggesting a common origin, while the other OTA PKSs analyzed were phylogenetically distant. A qRT-PCR assay monitored AcOTApks expression during fungal growth and concomitant production of OTA by A. carbonarius in synthetic grape medium. A clear correlation between the expression profile of AcOTApks and kinetics of OTA production was observed with

  14. Structure and stereospecificity of the dehydratase domain from the terminal module of the rifamycin polyketide synthase

    PubMed Central

    Gay, Darren; You, Young-Ok; Keatinge-Clay, Adrian; Cane, David E.

    2014-01-01

    RifDH10, the dehydratase domain from the terminal module of the rifamycin polyketide synthase, catalyzed the stereospecific syn dehydration of the model substrate (2S,3S)-2-methyl-3-hydroxypentanoyl-RifACP10, resulting in exclusive formation of (E)-2-methyl-2-pentenoyl-RifACP10. RifDH10 did not dehydrate any of the other three diastereomeric, RifACP10-bound, diketide thioester substrates. On the other hand, when EryACP6, from the sixth module of the erythromycin polyketide synthase, was substituted for RifACP10, RifDH10 stereospecifically dehydrated only (2R,3R)-2-methyl-3-hydroxypentanoyl-EryACP6 to give exclusively (E)-2-methyl-2-pentenoyl-EryACP6, with no detectable dehydration of any of the other three diastereomeric, EryACP6-bound, diketides. An identical alteration in substrate diastereospecificity was observed for the corresponding N-acetylcysteamine or pantetheine thioester analogues, regardless of acyl chain length or substitution pattern. Incubation of (2RS)-2-methyl-3-ketopentanoyl-RifACP10 with the didomain reductase-dehydratase RifKR10-RifDH10 yielded (E)-2-methyl-2-pentenoyl-RifACP10, the expected product of syn dehydration of (2S,3S)-2-methyl-3-hydroxypentanoyl-RifACP10, while incubation with the corresponding EryACP6-bound substrate, (2RS)-2-methyl-3-ketopentanoyl-EryACP6, gave only the reduction product (2S,3S)-2-methyl-3-hydroxypentanoyl-EryACP6 with no detectable dehydration. These results establish the intrinsic syn dehydration stereochemistry and substrate diastereoselectivity of RifDH10 and highlight the critical role of the natural RifACP10 domain in chaperoning the proper recognition and processing of the natural ACP-bound undecaketide substrate. The 1.82 Å-resolution structure of RifDH10 revealed the atomic resolution details of the active site and allowed modeling of the syn-dehydration of the (2S,3S)-2-methyl-3-hydroxyacyl-RifACP10 substrate. These results suggest that generation of the characteristic cis double bond of the rifamycins

  15. A 3-d modular gripper design tool

    SciTech Connect

    Brown, R.G.; Brost, R.C.

    1997-02-01

    Modular fixturing kits are sets of components used for flexible, rapid construction of fixtures. A modular vise is a parallel-jaw vise, each jaw of which is a modular fixture plate with a regular grid of precisely positioned holes. To fixture a part, one places pins in some of the holes so that when the vise is closed, the part is reliably located and completely constrained. The modular vise concept can be adapted easily to the design of modular parallel-jaw grippers for robots. By attaching a grid-plate to each jaw of a parallel-jaw gripper, one gains the ability to easily construct high-quality grasps for a wide variety of parts from a standard set of hardware. Wallack and Canny developed an algorithm for planning planar grasp configurations for the modular vise. In this paper, the authors expand this work to produce a 3-d fixture/gripper design tool. They describe several analyses they have added to the planar algorithm, including a 3-d grasp quality metric based on force information, 3-d geometric loading analysis, and inter-gripper interference analysis. Finally, the authors describe two applications of their code. One of these is an internal application at Sandia, while the other shows a potential use of the code for designing part of an agile assembly line.

  16. Evolution of Complex Modular Biological Networks

    PubMed Central

    Hintze, Arend; Adami, Christoph

    2008-01-01

    Biological networks have evolved to be highly functional within uncertain environments while remaining extremely adaptable. One of the main contributors to the robustness and evolvability of biological networks is believed to be their modularity of function, with modules defined as sets of genes that are strongly interconnected but whose function is separable from those of other modules. Here, we investigate the in silico evolution of modularity and robustness in complex artificial metabolic networks that encode an increasing amount of information about their environment while acquiring ubiquitous features of biological, social, and engineering networks, such as scale-free edge distribution, small-world property, and fault-tolerance. These networks evolve in environments that differ in their predictability, and allow us to study modularity from topological, information-theoretic, and gene-epistatic points of view using new tools that do not depend on any preconceived notion of modularity. We find that for our evolved complex networks as well as for the yeast protein–protein interaction network, synthetic lethal gene pairs consist mostly of redundant genes that lie close to each other and therefore within modules, while knockdown suppressor gene pairs are farther apart and often straddle modules, suggesting that knockdown rescue is mediated by alternative pathways or modules. The combination of network modularity tools together with genetic interaction data constitutes a powerful approach to study and dissect the role of modularity in the evolution and function of biological networks. PMID:18266463

  17. Managing in an age of modularity.

    PubMed

    Baldwin, C Y; Clark, K B

    1997-01-01

    Modularity is a familiar principle in the computer industry. Different companies can independently design and produce components, suck as disk drives or operating software, and those modules will fit together into a complex and smoothly functioning product because the module makers obey a given set of design rules. Modularity in manufacturing is already common in many companies. But now a number of them are beginning to extend the approach into the design of their products and services. Modularity in design should tremendously boost the rate of innovation in many industries as it did in the computer industry. As businesses as diverse as auto manufacturing and financial services move toward modular designs, the authors say, competitive dynamics will change enormously. No longer will assemblers control the final product: suppliers of key modules will gain leverage and even take on responsibility for design rules. Companies will compete either by specifying the dominant design rules (as Microsoft does) or by producing excellent modules (as disk drive maker Quantum does). Leaders in a modular industry will control less, so they will have to watch the competitive environment closely for opportunities to link up with other module makers. They will also need to know more: engineering details that seemed trivial at the corporate level may now play a large part in strategic decisions. Leaders will also become knowledge managers internally because they will need to coordinate the efforts of development groups in order to keep them focused on the modular strategies the company is pursuing.

  18. Theory for the Emergence of Modularity in Complex Systems

    NASA Astrophysics Data System (ADS)

    Deem, Michael; Park, Jeong-Man

    2013-03-01

    Biological systems are modular, and this modularity evolves over time and in different environments. A number of observations have been made of increased modularity in biological systems under increased environmental pressure. We here develop a theory for the dynamics of modularity in these systems. We find a principle of least action for the evolved modularity at long times. In addition, we find a fluctuation dissipation relation for the rate of change of modularity at short times. We discuss a number of biological and social systems that can be understood with this framework. The modularity of the protein-protein interaction network increases when yeast are exposed to heat shock, and the modularity of the protein-protein networks in both yeast and E. coli appears to have increased over evolutionary time. Food webs in low-energy, stressful environments are more modular than those in plentiful environments, arid ecologies are more modular during droughts, and foraging of sea otters is more modular when food is limiting. The modularity of social networks changes over time: stock brokers instant messaging networks are more modular under stressful market conditions, criminal networks are more modular under increased police pressure, and world trade network modularity has decreased

  19. Modular Wideband Active Vibration Absorber

    NASA Technical Reports Server (NTRS)

    Smith, David R.; Zewari, Wahid; Lee, Kenneth Y.

    1999-01-01

    A comparison of space experiments with previous missions shows a common theme. Some of the recent experiments are based on the scientific fundamentals of instruments of prior years. However, the main distinguishing characteristic is the embodiment of advances in engineering and manufacturing in order to extract clearer and sharper images and extend the limits of measurement. One area of importance to future missions is providing vibration free observation platforms at acceptable costs. It has been shown by researchers that vibration problems cannot be eliminated by passive isolation techniques alone. Therefore, various organizations have conducted research in the area of combining active and passive vibration control techniques. The essence of this paper is to present progress in what is believed to be a new concept in this arena. It is based on the notion that if one active element in a vibration transmission path can provide a reasonable vibration attenuation, two active elements in series may provide more control options and better results. The paper presents the functions of a modular split shaft linear actuator developed by NASA's Goddard Space Flight Center and University of Massachusetts Lowell. It discusses some of the control possibilities facilitated by the device. Some preliminary findings and problems are also discussed.

  20. The TOTEM modular trigger system

    NASA Astrophysics Data System (ADS)

    Bagliesi, M. G.; Berretti, M.; Cecchi, R.; Greco, V.; Lami, S.; Latino, G.; Oliveri, E.; Pedreschi, E.; Scribano, A.; Spinella, F.; Turini, N.

    2010-05-01

    The TOTEM experiment will measure the total cross-section with the luminosity independent method and study elastic and diffractive scattering at the LHC. We are developing a modular trigger system, based on programmable logic, that will select meaningful events within 2.5 μs. The trigger algorithm is based on a tree structure in order to obtain information compression. The trigger primitive is generated directly on the readout chip, VFAT, that has a specific fast output that gives low resolution hits information. In two of the TOTEM detectors, Roman Pots and T2, a coincidence chip will perform track recognition directly on the detector readout boards, while for T1 the hits are transferred from the VFATs to the trigger hardware. Starting from more than 2000 bits delivered by the detector electronics, we extract, in a first step, six trigger patterns of 32 LVDS signals each; we build, then, on a dedicated board, a 1-bit (L1) trigger signal for the TOTEM experiment and 16 trigger bits to the CMS experiment global trigger system for future common data taking.