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The biosynthesis of the ployketide knipholone, as an axially chiral phenylanthraquinone, in higher plants is examined by feeding the plant with labeled precursors [super]13[C.sub.2]- acetate and other precursors. It is found that, based on the biosynthetic origin of knipholone, the northern portion of the molecule, the anthraquinone moiety, is built up from eight acetate units according to the folding mode F and the southern part built from four acetate units is localized, not [C.sub.2]-wise randomized.

An attempt is made to reconcile some of the inconsistencies identified in a collection of proteins to selectively recognize di-and trimethyl H3K4, marks that are associated with actively transcribed genes. It is revealed that factors known to be involved with active transcription associate with H3K4me2/3 has important significance for understanding the functional role of this histone modification.

Combinatorial biosynthesis of type 1 polyketide synthases is a promising approach for the generation of new structural derivatives of polyketide-containing natural products. A target of this approach has been to change the extender units incorporated into a polyketide backbone to alter the structure and activity of the natural product. One limitation to these efforts is that only four extender units were known: malonyl-CoA, methylmalonyl-CoA, ethylmalonyl-CoA, and methoxymalonyl-acyl carrier protein (ACP). The chemical attributes of these extender units are quite similar, with the exception of the potential hydrogen bonding interactions by the oxygen of the methoxy moiety. Furthermore, the incorporated extender units are not easily modified by using simple chemical approaches when combinatorial biosynthesis is coupled to semisynthetic chemistry. We recently proposed the existence of two additional extender units, hydroxymalonyl-ACP and aminomalonyl-ACP, involved in the biosynthesis of zwittermicin A. These extender units offer unique possibilities for combinatorial biosynthesis and semisynthetic chemistry because of the introduction of free hydroxyl and amino moieties into a polyketide structure. Here, we present the biochemical and mass spectral evidence for the formation of these extender units. This evidence shows the formation of ACP-linked extender units for polyketide synthesis. Interestingly, aminomalonyl-ACP formation involves enzymology typically found in nonribosomal peptide synthesis. antibiotics | combinatorial biosynthesis

Trianglelac-acetogenins, a new class of inhibitors of bovine heart mitochondrial complex I (NADH-ubiquinone oxidoreductase) act differently from ordinary inhibitors such as rotenone and piericidin A. The results show that binding site of trianglelac-acetogenins is not identical to the binding site of ubiquinone and downstream of that of ordinary inhibitors.

The biosynthetic gene cluster for the aromatic polyketide steffimycin of the anthracycline family was cloned and characterized from 'Streptomyces steffisburgensis' NRRL 3193. Sequence analysis of a 42.8-kbp DNA region revealed the 36 open reading frames (ORFs) (one of them incomplete), 24 of which spanning 26.5 kb, are probably involved in steffimycin biosynthesis.

The formation of a carbon-carbon bond is an essential step in the biosynthetic pathways by which fatty acids and polyketides are made. The thiolase superfamily enzymes catalyse this carbon-carbon-bond formation via a thioester-dependent Claisen-condensation-reaction mechanism. In this way, fatty-acid chains and polyketides are made by sequentially adding simple building blocks, such as acetate units, to the growing molecule. A common feature of these enzymes is a reactive cysteine residue that is transiently acylated in the catalytic cycle. The wide catalytic diversity of the thiolase superfamily enzymes is of great interest. In particular, the type-III polyketide synthases make complicated compounds of great biological importance using multiple, subsequent condensation reactions, which are all catalysed in the same active-site cavity. The crucial metabolic importance of the bacterial fatty-acid-synthesizing enzymes stimulates in-depth studies that aim to develop efficient anti-bacterial drugs.

Studies were conducted on the inhibitory mechanism of acetogenins, the most potent inhibitors of mitochondrial complex I (NADH-ubiquinone oxidoreductase, which are useful for explaining the structural and functional feature of the terminal electron transfer step of enzyme. It suggested that the spacer portion is free from steric congestion arising from the putative binding site probably because there is no cavity-like binding site for the spacer portion.

A novel polyketide metabolite, thailandamide lactone, which is not detected in the wild type, is isolated from the mutant broth, and its structure is analyzed by high-resolution mass spectrometry and IR and NMR spectroscopy. In a biological assay tumor cell lines, thialandamide lactone has displayed moderate antiproliferative activities.

The complete functions of Aspergillus nidulans aspyridone synthetase (ApdA) and its enoylreductase partner ApdC are restructured in vitro and in Saccharomyces cerevisiae with the production of preaspyridone and the programming rules of polyketide synthase (PKS) and nonribosomal peptide synthetase (NRPS) modules are examined in vitro. The results have represented one of the largest, multidomain enzymes and have provided new opportunities for engineered biosynthesis of fungal natural products.

A method is described by combining chemobiosynthesis and engineered minipolyketide synthases (PKSs) that has produced polyketide intermediates possessing multiple stereogenic centers. The method is validated by generating the intermediates in Escherichia coli for the synthesis of trans-9,10-dehydroepothilone D.

Cloning, sequencing, heterologous expression, and mechanistic analysis of A-74528 biosynthesis, natural product of Streptomyces sp. which inhibits a key regulatory enzyme 2',5'-oligoadenylate phosphodiesterase (2'-PDE) of the interferon pathway is demonstrated. The gene A-74528 which is found to possess a type II polyketide synthase (PKS) at its core could have significant implications for the evolution of antibiotics with different biological activities from the same gene cluster.

The initiation polyketide synthase (PKS) module responsible for hexadienyl priming of the antibiotic fredericamycin was identified to study its biochemical properties. The utility of the PKS module to the metabolic engineer is demonstrated by the design and in vivo biosynthesis of unusually primed analogues of a representative polyketide product.

The article discusses a recently devised reaction sequence that can be employed for the stereochemical synthesis of various spiroketals. The mechanisms associated with the polyketide assembly taking place via the alkene-alkyne cross-coupling reaction are also analyzed.

Structural analysis and genetic and biochemical experiments were performed for BE-7585A produced by Amycolatopsis orientalis subsp. vinearia BA-07585 which has a 2-thiosugar and is a member of the angucycline class of compounds. The spectroscopic analyses verified the structure of BE-7585A to be closely related to rhodonocardin A and provided insight into the biosynthetic pathway for BE-7585A which could be applied for future biochemical studies of 2-thiosugar biosynthesis and BE-7585A assembly.

The synthesis and characterization of new analogues resulting from inactivation of two tautomycetin (TTN) genes, ttnD and ttnF, in Streptomyces griseochromogenes are described. The results provide better insight into the transformations responsible for converting the post-polyketide synthase product into the exciting anticancer and immunosuppressive chemotherapeutic candidate TTN and significance of functional groups installed by TtnDF.

A convergent strategy with longest linear sequence of 23 steps was used to achieve a highly stereoselective joint total synthesis of the potent polyketide macrolide antibiotics etnangien and etnangien methyl ester. The expedient and flexible strategy of the etnangiens could be adapted to design analogues of these RNA-polymerase inhibitors and allow further exploration of the promising biological potential of these macrolide antibiotics.

The role of natural products in the history of organic chemistry and the biosynthesis of polyketides and non-ribosomal peptides are discussed. The study has analyzed the monomer biogenesis, coupling chemistries from templated and non-templated pathways and the broad set of tailoring reactions and hybrid pathways giving rise to the diverse scaffolds and functionalization patterns of natural products.

Photolysis reaction of 1-methyl-9,10-anthraquinones is performed in the presence of oxygen to obtain endoperoxides that could be reduced to produce 1-hydroxymethyl-9,10-anthraquinones. The photochemical hydroxylation could also be used for total synthesis of a naturally occurring polyketide, 9'-hydroxyaloesaponarin II.

The biochemical basis for controlling polyketide stereochemistry is examined and the timing and mechanism of the epimerization at methyl-bearing centers is established by carrying out a series of incubations by using reconstituted components from modular polyketide synthases. The studies have shown that the stereochemistry of the product is directly correlated with the intrinsic stereospecificity of the ketoreductase domain and the epimerization is catalyzed by the ketoreductase domain itself.

A highly stereoselective formal total synthesis of the ornithine decarboxylase inhibitors (-)-saliniketals A and B is described. The salient features in the synthesis involved desymmetrization technique to create six contiguous chiral centers from a single bicyclic precursor as well as substrate-controlled Grignard reaction, intramolecular Wacker-type oxidation, and antialdol reaction following Pirrung-Heathcock conditions.

An unprecedented decarboxylative chain termination mechanism which could be used for the polyketide synthase of curacin A, an anticancer lead compound isolated from the marine cyanobacterium Lyngbya majuscula is described. The biochemical characterization of the unusual chain termination module containing adjacent sulfotransferase (ST) and thioesterase (TE) catalytic domains embedded in CurM which could be applied to obtain rare terminal olefin is also described.

The mechanisms involved in the synthesis of the ABC-rings of the polyketide natural product hexacyclinic acid are discussed. The different intermediates and processes used for the development of different rings of the acid are also explained.

A longest sequence of 21 steps, without resorting to stoichiometric chiral auxiliaries or the chiral pool was utilized to perform catalytic enantioselective synthesis of the antimelanoma marine natural product (-)-palmerolide A. The unique method could also open new pathways for the design of simplified analogues of palmerolide.

Several nuclear magnetic resonance (NMR) analyses are conducted to explain the synthesis of a series of the C1-C17 fragments of a new natural polyketide. The relative stereochemistry of the fragment of polyketide portion is also explained.

The simultaneous in vitro production of three octaketide polyenes by biosynthetic enzymes for the 10-membered enediyne calicheamicin [[gamma].sub.1.sup.1], including the elusive [beta]-keto acid precursor to C15 methyl hexaenone, is described. The synthesis of this set of octaketides by the enediyne polyketide synthases (PKS) CalE8 and thioesterase CalE7 has shown that accessory enzymes are required for initiating differentiation to 9- or 10-membered enediyne precursors.

The structures and the mechanisms of the two polyketide synthases (PKSs) are discussed.

The full length LovF is expressed and has reconstituted the megasynthase activities in vitro. The catalytic efficiency is increased when LovF, a highly reducing polyketide synthase (HR-PKS) from the filamentous fungus Aspergillus terreus, is used as the acyl carrier instead of N-acetylcysteamine.

The acyl carrier protein (ACP) of FUM1 and in vitro loaded acyl chains are expressed to the ACP from acyl-CoA by using a 4'-phosphopantetheinyl transferase. The analysis of mycotoxin fumonisins has shown a novel polyketide chain-releasing mechanism, in which a pyridoxal 5'-phosphate (PLP)-dependent enzyme has catalyzed the termination and offloading of the polyketide chains as well as the introduction of a new carbon-carbon bond and an amino group to the chain.

The construction of hydrindane ring system of coronafacic acid is examined in order to show that the canonical polyketide chain elongation and functionalization encoded by type I modular polyketide synthases (PKSs) has taken place along with cascade reactions during assembly line-like biosynthesis. The latent reactivity within the substrate is disclosed upon elongation in order for cascade reactions to proceed at a rate faster than the functionalization reactions encoded by the catalytic domains within the type I PKS.

The formation of unsaturated lactone in the phoslactomycins (PLMs) and other related compounds are shown to be dependent upon an enzyme such as Plm[T.sub.2], not a canonical dehydratase (DH) domain in the modular polyketide synthase (PKS). The M-PLM B has undergone rapid elimination to generate the cis [[DELTA].sup.2,3] alkene of the PLMs in a process that might reflect that catalyzed by Plm[T.sub.2].

The first synthesis and characterization of a diverse natural product with complex core structure such as iso-9,10-deoxytridachione is reported. The results obtained for products derived from strained all-(E) tetracene-pyrone precursor provide insight into the biomimetic transformation under a variety of reaction conditions.

A convenient approach is developed for the synthesis of 10-deoxymethynolide and narbonolide, which as aglycones of the methymycin and the pikromycin families of macrolide antibiotics. The ring-closing metathesis reaction catalyzed by the second-generation Grubbs catalyst is efficient in preparing the macrocyclic polyketide lactones.

The preparation and structural studies of (+)-okilactomycin, a novel polyketide antitumor antibiotic isolated from a bioactive filtrate of Streptomyces griseoflavus is reported.

The study describes a new mechanism for the formation of benzopyrone in aromatic polyketide biosynthesis, which otherwise is present in numerous natural products. Benzopyrones are normally formed through the esterification of carboxylic acids and intramolecular hydrolysis of thioesters.

CmaE is a commonly used transferase, which helps in moving the aminoacyl groups between various carrier protein domains in the coronamic acid biosynthetic pathway. The analysis shows that CmaE can also be used for generating chemical diversity within a nonribosomalpeptide (NRPS).

The formation of a nonaketide by the structurally simple type III polyketide synthase (PKS) is presented. The structure-based engineering of the plant-specific chalcone synthase (CHS) superfamily type III PKS enzymes will produce the chemically and structurally disparate unnatural novel polyketides.

The stereochemistry of the polyketide intermediate, which is transferred from module 1 to module 2 of phoslactomycins (PLM), is determined to distinguish between the possibilities of establishing the trans and cis double bond directly. The results have shown that only the N-acetylcysteamine derivative of the cis-diketide can prime PLM2 directly and that other diketides produces PLM B production only through degradation to an activated cyclohexanecarboxylic acid and elongation using PLM1.

The characterization of a stereoisomer library of 28 of the 64 possible isomers of the acetogenin murisolin including 24 of the 32 possible diastereomers has provided a complete picture of the spectra of this class of molecules. The structure of murisolin is proved to be the 4R,15R,16R,19R,20R,34R isomer, whereas the assignment of 16,19-cis-murisolin as RRRSSS is changed to the RSSRRS diastereomer and murisolin A is suggested to be RSRRRS.

The concept of molecular simplification as a drug design strategy to shorten synthetic routes, while keeping or enhancing the biological activity of the lead drug, is applied to (+)-muconin, an acetogenin with remarkable cytotoxicity. A novel method that enables the stereoselective synthesis of such a natural compound or its enantiomer from a common precursor, affording complete stereochemical control and decrease in the number of chemical steps required, is described.

The extender unit and acyl carrier protein (ACP) specialty of keto synthases (KS) domains from modules 3 and 6 of 6-deoxyerythronolide B synthase (DEBS) are analyzed to understand the engineering opportunities of these assembly lines. A new method for engineering polyketide synthase (PKS) modules is described that is based upon kinetic analysis of the substrate and ACP preferences of individual KS domains, as well as the affect of interdomain linkers within a PKS module.

The characterization of FUM14 (where FUM = 15-gene cluster) helps in revealing the biosynthetic mechanism for the side-chain formation of fumonisins and also in understanding the reactions catalyzed by nonribosomal peptide synthetase (NRPS), resulting in the formation of tricarballylic esters. The gene disruption and heterologous expression of FUM14 and in vitro activity assays for the purified Fum14p (FUM14 protein) are reported.

An enantioselective total synthesis of the proposed structure of mucoxin, which features two regio- and stereoselective tetrahydrofuran (THF) ring-forming reactions, is described. The spectroscopic analysis of the synthesized molecule shows that the spectral discrepancies are due to stereochemical misassignment of the natural product.

The potent biological activity and structural diversity of the polyketide class of natural products are studied. In the preliminary survey of structural generality ketene acetals bearing methyl groups on the alpha and beta atoms of the dienyl unit are prepared.

The synthesis of simple diketide N-acetylcysteamine (SNAC)-seco-10-deoxymethynolide, a mimic of the natural hexaketide Pik thioesterase (TE) chain elongation intermediate, and its enzymatic macrolactonization by Pik TE to complete the total synthesis of 10-deoxymetynolide is reported. The exquisite selectivity and efficiency of this enzyme is revealed.

The natural pentaketide and hexaketide chain elongation intermediates as N-acetyl cysteamine (NAC) thioesters is synthesized. These are used as substrates for in vitro conversion with engineered PikAIII+TE and in combination with native PikIII (module 5) and PikAIV (module 6) multifunctional proteins.

The expression and purification of recombinant FkbP from the FK520 biosynthetic pathway is reported. The observations suggest that the adenylation domain of FkbP serves as the primary selectivity filter for pipecolate incorporation.

Muconin is a sequential tetrahydrofuran/tetrahydropuran (THF/THP)-processing acetogenin that exhibits potent and selective in vitro cytotoxicity toward pancreatic and breast tumor cell lines. A new route is established for obtaining (+)-muconin starting with (-)-muricatacin, a compound synthesized via the novel alpha-C-H hydroxyalkylation and alpha'-C-H oxidation of tetrahydrofuran.

Two strategies are described for redirecting the cyclization steps of fungal polyketide synthases (PKSs). The cooperative activities of fungal and bacterial PKS components are very important and have enabled the synthesis of polyketides using enzymes from two distinct families of PKSs.