Genome-Scale Model of Rhizopus microsporus: Metabolic integration of a fungal holobiont with its bacterial and viral endosymbionts.

Rhizopus microsporus often lives in association with bacterial and viral symbionts that alter its biology. This fungal model represents an example of the complex interactions established among diverse organisms in functional holobionts. We constructed a Genome-Scale Model (GSM) of the fungal-bacterial-viral holobiont (iHol). We employed a constraint-based method to calculate the metabolic fluxes to decipher the metabolic interactions of the symbionts with their host. Our computational analyses of iHol simulate the holobiont's growth and the production of the toxin rhizoxin. Analyses of the calculated fluxes between R. microsporus in symbiotic (iHol) versus asymbiotic conditions suggest that changes in the lipid and nucleotide metabolism of the host are necessary for the functionality of the holobiont. Glycerol plays a pivotal role in the fungal-bacterial metabolic interaction, as its production does not compromise fungal growth, and Mycetohabitans bacteria can efficiently consume it. Narnavirus RmNV-20S and RmNV-23S affected the nucleotide metabolism without impacting the fungal-bacterial symbiosis. Our analyses highlighted the metabolic stability of Mycetohabitans throughout its co-evolution with the fungal host. We also predicted changes in reactions of the bacterial metabolism required for the active production of rhizoxin. This iHol is the first GSM of a fungal holobiont.

Cyanobacteria: A Promising Source of Antifungal Metabolites.

Cyanobacteria are a rich source of secondary metabolites, and they have received a great deal of attention due to their applicability in different industrial sectors. Some of these substances are known for their notorious ability to inhibit fungal growth. Such metabolites are very chemically and biologically diverse. They can belong to different chemical classes, including peptides, fatty acids, alkaloids, polyketides, and macrolides. Moreover, they can also target different cell components. Filamentous cyanobacteria have been the main source of these compounds. This review aims to identify the key features of these antifungal agents, as well as the sources from which they are obtained, their major targets, and the environmental factors involved when they are being produced. For the preparation of this work, a total of 642 documents dating from 1980 to 2022 were consulted, including patents, original research, review articles, and theses.

Marine Cyanobacteria as Sources of Lead Anticancer Compounds: A Review of Families of Metabolites with Cytotoxic, Antiproliferative, and Antineoplastic Effects.

The marine environment is highly diverse, each living creature fighting to establish and proliferate. Among marine organisms, cyanobacteria are astounding secondary metabolite producers representing a wonderful source of biologically active molecules aimed to communicate, defend from predators, or compete. Studies on these molecules' origins and activities have been systematic, although much is still to be discovered. Their broad chemical diversity results from integrating peptide and polyketide synthetases and synthases, along with cascades of biosynthetic transformations resulting in new chemical structures. Cyanobacteria are glycolipid, macrolide, peptide, and polyketide producers, and to date, hundreds of these molecules have been isolated and tested. Many of these compounds have demonstrated important bioactivities such as cytotoxicity, antineoplastic, and antiproliferative activity with potential pharmacological uses. Some are currently under clinical investigation. Additionally, conventional chemotherapeutic treatments include drugs with a well-known range of side effects, making anticancer drug research from new sources, such as marine cyanobacteria, necessary. This review is focused on the anticancer bioactivities of metabolites produced by marine cyanobacteria, emphasizing the identification of each variant of the metabolite family, their chemical structures, and the mechanisms of action underlying their biological and pharmacological activities.

Systems metabolic engineering of Streptomyces venezuelae for the enhanced production of pikromycin.

Pikromycin is an important precursor of drugs, for example, erythromycin. Hence, systems metabolic engineering for the enhanced pikromycin production can contribute to the development of pikromycin-related drugs. In this study, metabolic genes in Streptomyces venezuelae were systematically engineered for enhanced pikromycin production. For this, a genome-scale metabolic model of S. venezuelae was reconstructed and simulated, which led to the selection of 11 metabolic gene targets. These metabolic genes, including four overexpression targets and seven knockdown targets, were individually engineered first. Next, two overexpression targets and two knockdown targets were selected based on the 11 strains' production performances to engineer two to four of these genes together for the potential synergistic effects on the pikromycin production. As a result, the NM1 strain with AQF52_RS24510 (methenyltetrahydrofolate cyclohydrolase/methylenetetrahydrofolate dehydrogenase) overexpression and AQF52_RS30320 (sulfite reductase) knockdown showed the best production performance among all the 22 strains constructed in this study. Fed-batch fermentation of the NM1 strain produced 295.25 mg/L of pikromycin, by far the best production titer using the native producer S. venezuelae, to the best of our knowledge. The systems metabolic engineering strategy demonstrated herein can also be applied to the overproduction of other secondary metabolites using S. venezuelae.

In vitro biotransformation of OA-group and PTX-group toxins in visceral and non-visceral tissues of Mytilus chilensis and Ameghinomya antiqua.

Lipophilic marine toxins (LMTs) are made up of multiple groups of toxic analogues, which are characterised by different levels of cellular and toxic action. The most prevalent groups in the southern Pacific zone are: a) okadaic acid group (OA-group) which consists of okadaic acid (OA) and dinophysistoxin-1 (DTX-1); and, b) pectenotoxin-2 (PTX2) group which consists of pectenotoxin-2 (PTX-2). The main objective of our study was to examine in vitro biotransformation of OA-group and PTX-group in the tissues of two endemic species of bivalves from southern Chile; blue mussels (Mytilus chilensis) and clams (Ameghinomya antiqua). The biotransformation processes of both groups were only detected in the digestive glands of both species using LC-MS/MS. The most frequently detected analogues were acyl derivatives (≈2.0 ± 0.1 µg ml-1) for OA-group and PTX-2SA (≈1.4 ± 0.1 µg ml-1) for PTX-group, with a higher percentage of biotransformation for OA-group (p < .001). In addition, simultaneous incubations of the different analogues (OA/PTX-2; DTX-1/PTX-2 and OA/DTX-1/PTX-2) did not show any interaction between the biotransformation processes. These results show that the toxicological variability of endemic species leads to biotransformation of the profile of toxins, so that these new analogues may affect people's health.

Aerobic dissipation of avermectins and moxidectin in subtropical soils and dissipation of abamectin in a field study.

Avermectins and moxidectin are antiparasitics widely used as active pharmaceutical ingredients in veterinary medicine, as well as in pesticide formulations for pest control in agriculture. Although the use of these compounds provides benefits to agribusiness, they can impact the environment, since a large part of these substances may reach the soil and water from the excreta of treated animals and following direct applications to crops. The present work had the objective of evaluating the dissipation behaviors of abamectin, doramectin, eprinomectin, ivermectin, and moxidectin in four native Brazilian soils of different textural classes (clay, sandy-clay, sandy, and sandy-clay-loam), following OECD Guideline 307. The studies were conducted in a climate chamber at 22 °C, 71% relative humidity, and protected from light. The dissipation studies were carried out with all drugs together, since no difference was verified when studies were done with each drug separately. The concentrations of the drugs in the soils were determined using an ultra-high performance liquid chromatograph coupled to a fluorescence detector or a tandem mass spectrometer. The dissipation half-life (DT50) values ranged from 9 to 16 days and the calculated GUS index values were in the range from -1.10 to 0.08, indicating low mobility of the drugs in the soils evaluated and low tendency for leaching. In addition, a field study was carried out to evaluate the dissipation of abamectin after application of a foliar pesticide in an orange crop. A DT50 of 9 days was determined, which was similar to that obtained under controlled conditions in the climate chamber (12 days), indicating that biotransformation was the primary process influencing the overall dissipation.

Growth and differentiation properties of pikromycin-producing Streptomyces venezuelae ATCC15439.

Streptomycetes naturally produce a variety of secondary metabolites, in the process of physiological differentiation. Streptomyces venezuelae differentiates into spores in liquid media, serving as a good model system for differentiation and a host for exogenous gene expression. Here, we report the growth and differentiation properties of S. venezuelae ATCC-15439 in liquid medium, which produces pikromycin, along with genome-wide gene expression profile. Comparison of growth properties on two media (SPA, MYM) revealed that the stationary phase cell viability rapidly decreased in SPA. Submerged spores showed partial resistance to lysozyme and heat, similar to what has been observed for better-characterized S. venezuelae ATCC10712, a chloramphenicol producer. TEM revealed that the differentiated cells in the submerged culture showed larger cell size, thinner cell wall than the aerial spores. We analyzed transcriptome profiles of cells grown in liquid MYM at various growth phases. During transition and/or stationary phases, many differentiationrelated genes were well expressed as judged by RNA level, except some genes forming hydrophobic coats in aerial mycelium. Since submerged spores showed thin cell wall and partial resistance to stresses, we examined cellular expression of MreB protein, an actin-like protein known to be required for spore wall synthesis in Streptomycetes. In contrast to aerial spores where MreB was localized in septa and spore cell wall, submerged spores showed no detectable signal. Therefore, even though the mreB transcripts are abundant in liquid medium, its protein level and/or its interaction with spore wall synthetic complex appear impaired, causing thinner- walled and less sturdy spores in liquid culture.

Production of pikromycin using branched chain amino acid catabolism in Streptomyces venezuelae ATCC 15439.

Branched chain amino acids (BCAA) are catabolized into various acyl-CoA compounds, which are key precursors used in polyketide productions. Because of that, BCAA catabolism needs fine tuning of flux balances for enhancing the production of polyketide antibiotics. To enhance BCAA catabolism for pikromycin production in Streptomyces venezuelae ATCC 15439, three key enzymes of BCAA catabolism, 3-ketoacyl acyl carrier protein synthase III, acyl-CoA dehydrogenase, and branched chain α-keto acid dehydrogenase (BCDH) were manipulated. BCDH overexpression in the wild type strain resulted in 1.3 fold increase in pikromycin production compared to that of WT, resulting in total 25 mg/L of pikromycin. To further increase pikromycin production, methylmalonyl-CoA mutase linked to succinyl-CoA production was overexpressed along with BCDH. Overexpression of the two enzymes resulted in the highest titer of total macrolide production of 43 mg/L, which was about 2.2 fold increase compared to that of the WT. However, it accumulated and produced dehydroxylated forms of pikromycin and methymycin, including their derivatives as well. It indicated that activities of pikC, P450 monooxygenase, newly became a bottleneck in pikromycin synthesis.

Modulation of lateral and longitudinal interdimeric interactions in microtubule models by Laulimalide and Peloruside A association: A molecular modeling approach on the mechanism of microtubule stabilizing agents.

Laulimalide (LAU) and Peloruside A (PLA) are non-taxane microtubule stabilizing agents with promising antimitotic properties. These ligands promote the assembly of microtubules (MTs) by targeting a unique binding site on ß-tubulin. The X-ray structure for LAU/PLA-tubulin association was recently elucidated, but little information is available regarding the role of these ligands as modulators of interdimeric interactions across MTs. Herein, we report the use of molecular dynamics (MD), principal component analysis (PCA), MM/GBSA-binding free energy calculations, and computational alanine scanning mutagenesis (ASM) to examine effect of LAU/PLA association on lateral and longitudinal contacts between tubulin dimers in reduced MT models. MD and PCA results revealed that LAU/PLA exerts a strong restriction of lateral and longitudinal interdimeric motions, thus enabling the stabilization of the MT lattice. Besides structural effects, LAU/PLA induces a substantial strengthening of longitudinal interdimeric interactions, whereas lateral contacts are less affected by these ligands, as revealed by MM/GBSA and ASM calculations. These results are valuable to increase understanding about the molecular features involved in MT stabilization by LAU/PLA, and to design novel compounds capable of emulating the mode of action of these ligands.

Target-specific identification and characterization of the putative gene cluster for brasilinolide biosynthesis revealing the mechanistic insights and combinatorial synthetic utility of 2-deoxy-l-fucose biosynthetic enzymes.

Brasilinolides exhibiting potent immunosuppressive and antifungal activities with remarkably low toxicity are structurally characterized by an unusual modified 2-deoxy-l-fucose (2dF) attached to a type I polyketide (PK-I) macrolactone. From the pathogenic producer Nocardia terpenica (Nocardia brasiliensis IFM-0406), a 210 kb genomic fragment was identified by target-specific degenerate primers and subsequently sequenced, revealing a giant nbr gene cluster harboring genes (nbrCDEF) required for TDP-2dF biosynthesis and those for PK-I biosynthesis, modification and regulation. The results showed that the genetic and domain arrangements of nbr PK-I synthases agreed colinearly with the PK-I structures of brasilinolides. Subsequent heterologous expression of nbrCDEF in Escherichia coli accomplished in vitro reconstitution of TDP-2dF biosynthesis. The catalytic functions and mechanisms of NbrCDEF enzymes were further characterized by systematic mix-and-match experiments. The enzymes were revealed to display remarkable substrate and partner promiscuity, leading to the establishment of in vitro hybrid deoxysugar biosynthetic pathways throughout an in situ one-pot (iSOP) method. This study represents the first demonstration of TDP-2dF biosynthesis at the enzyme and molecular levels, and provides new hope for expanding the structural diversity of brasilinolides by combinatorial biosynthesis.

Predictive criteria of insignificant prostate cancer: what is the correspondence of linear extent to percentage of cancer in a single core?

Objective The aim of active surveillance of early prostate cancer is to individualize therapy by selecting for curative treatment only patients with significant cancer. Epstein’s criteria for prediction of clinically insignificant cancer in surgical specimens are widely used. Epstein’s criterion “no single core with >50% cancer” has no correspondence in linear extent. The aim of this study is to find a possible correspondence. Materials and Methods From a total of 401 consecutive patients submitted to radical prostatectomy, 17 (4.2%) met criteria for insignificant cancer in the surgical specimen. The clinicopathologic findings in the correspondent biopsies were compared with Epstein’s criteria for insignificant cancer. Cancer in a single core was evaluated in percentage as well as linear extent in mm. Results Comparing the clinicopathologic findings with Epstein’s criteria predictive of insignificant cancer, there was 100% concordance for clinical stage T1c, no Gleason pattern 4 or 5, ≤2 cores with cancer, and no single core with >50% cancer. However, only 25% had density ≤0.15. The mean, median and range of the maximum length of cancer in a single core in mm were 1.19, 1, and 0.5-2.5, respectively. Additionally, the mean, median, and range of length of cancer in all cores in mm were 1.47, 1.5, and 0.5-3, respectively. Conclusion To pathologists that use Epstein’s criteria predictive of insignificant cancer and measure linear extent in mm, our study favors that “no single core with >50% cancer” may correspond to >2.5 mm in linear extent. .

Serum insulin-like growth factor-1 levels in females and males in different cervical vertebral maturation stages

OBJECTIVE: The aim of this cross sectional study was to assess serum insulin-like growth factor-1 (IGF-1) levels in female and male subjects at various cervical vertebral maturation (CVM) stages. MATERIAL AND METHODS: The study sample consisted of 60 subjects, 30 females and 30 males, in the age range of 8-23 years. For all subjects, serum IGF-1 level was estimated from blood samples by means of chemiluminescence immunoassay (CLIA). CVM was assessed on lateral cephalograms using the method described by Baccetti. Serum IGF-1 level and cervical staging data of 30 female subjects were included and taken from records of a previous study. Data were analyzed by Kruska-Wallis and Mann Whitney test. Bonferroni correction was carried out and alpha value was set at 0.003. RESULTS: Peak value of serum IGF-1 was observed in cervical stages CS3 in females and CS4 in males. Differences between males and females were observed in mean values of IGF-1 at stages CS3, 4 and 5. The highest mean IGF-1 levels in males was observed in CS4 followed by CS5 and third highest in CS3; whereas in females the highest mean IGF-1 levelswas observed in CS3 followed by CS4 and third highest in CS5. Trends of IGF-1 in relation to the cervical stages also differed between males and females. The greatest mean serum IGF-1 value for both sexes was comparable, for females (397 ng/ml) values were slightly higher than in males (394.8 ng/ml). CONCLUSIONS: Males and females showed differences in IGF-1 trends and levels at different cervical stages. .

OBJETIVO: o objetivo do presente estudo transversal foi avaliar os níveis do fator de crescimento semelhante à insulina-1 (IGF-1 sérico) em pacientes de ambos os sexos e em diferentes estágios de maturação das vértebras cervicais (MVC). MÉTODOS: a amostra consistiu de 60 pacientes, sendo 30 do sexo masculino e 30 do sexo feminino, com idades entre 8 e 23 anos. Amostras de sangue foram colhidas de todos os pacientes, cujos níveis de IGF-1 sérico foram avaliados por meio do método de imunoensaio quimioluminescente (CLIA). O estágio de MVC foi avaliado por meio de radiografias cefalométricas de perfil por meio do método descrito por Baccetti. O nível de IGF-1 sérico e o estágio de maturação das vertebras cervicais de 30 pacientes do sexo feminino foram avaliados e os dados retirados dos registros de um estudo prévio. Os dados foram submetidos aos testes de Kruskal-Wallis e de Mann-Whitney. A correção de Bonferroni foi calculada e o valor de alfa foi de 0,003. RESULTADOS: o valor de pico do IGF-1 sérico foi encontrado no estágio CS3, para mulheres, e CS4, para homens. Foram encontradas diferenças entre as médias dos valores de IGF-1 entre homens e mulheres nos estágios CS3, 4 e 5. O valor médio mais alto para os níveis de IGF-1 nos homens foi observado no estágio CS4, seguido do estágio CS5 e CS3. Nas mulheres, o valor médio mais alto foi observado em CS3, seguido do estágio CS4 e CS5. Diferenças também foram encontradas quanto à curva do IGF-1, em relação ao estágio de maturação das vértebras cervicais nos pacientes de ambos os sexos. O valor médio de IGF-1 sérico mais alto foi comparado. As pacientes do sexo feminino apresentaram valores ligeiramente mais altos (397ng/ml) em comparação aos pacientes do sexo masculino (394.8ng/ml). CONCLUSÕES: homens e mulheres apresentam valores de IGF-1 diferentes em estágios de maturação das vértebras cervicais diferentes. .

Effects of Sucrose, Phosphate, and Calcium Carbonate on the Production of Pikromycin from Streptomyces venezuelae.

Polyketide secondary metabolites share common precursor pools, acyl-CoA. Thus, the effects of engineering strategies for heterologous and native secondary metabolite production are often determined by the measurement of pikromycin in Streptomyces venezuelae. It is hard to compare the effectiveness of engineering targets among published data owing to the different pikromycin production media used from one study to the other. To determine the most important nutritional factor and establish optimal culture conditions, medium optimization of pikromycin from Streptomyces venezuelae ATCC 15439 was studied with a statistical method, Plackett-Burman design. Nine variables (glucose, sucrose, peptone, (NH4)2SO4, K2HPO4, KH2PO4, NaCl, MgSO4·7H2O, and CaCO3) were analyzed for their effects on a response, pikromycin. Glucose, K2HPO4, and CaCO3 were determined to be the most significant factors. The path of the steepest ascent and response surface methodology about the three selected components were performed to study interactions among the three factors, and the fine-tune concentrations for maximized product yields. The significant variables and optimal concentrations were 139 g/1 sucrose, 5.29 g/l K2HPO4, and 0.081 g/l CaCO3, with the maximal pikromycin yield of 35.5 mg/l. Increases of the antibiotics production by 1.45-fold, 1.3-fold, and 1.98-fold, compared with unoptimized medium and two other pikromycin production media SCM and SGGP, respectively, were achieved.

[Paradoxical reactions and responses during antibiotic treatment for Mycobacterium ulcerans infection (Buruli ulcer). Four cases from French Guiana]. / Réponses et réactions paradoxales au cours du traitement médicamenteux de l'infection à Mycobacterium ulcerans (ulcère de Buruli). Quatre observations en Guyane française.

BACKGROUND: In recent years, first-line therapy for Mycobacterium ulcerans infection in French Guiana has consisted of antibiotics active against this organism. Two regimens are used comprising rifampicin associated with clarithromycin or amikacin. PATIENTS AND METHODS: We describe four patients presenting apparent worsening of their lesions during treatment: ulceration of a nodular lesion in a 32-year-old woman and worsening of an ulcerated lesion in three patients aged 16, 27 and 79 years. DISCUSSION: In these 4 patients, we concluded that the symptoms were caused by a paradoxical response or a reaction, a phenomenon already described in tuberculosis and leprosy. Such worsening is transient and must not be misinterpreted as failure to respond to treatment. The most plausible pathophysiological hypothesis involves the re-emergence of potentially necrotizing cellular immunity secondary to the loss of mycolactone, a necrotizing and immunosuppressive toxin produced by M. ulcerans, resulting from the action of the antibiotics.

Isolation and characterization of phytotoxic compounds produced by Streptomyces sp. AMC 23 from red mangrove (Rhizophora mangle).

Natural products produced by microorganisms have been utilized as sources of new drugs possessing a wide range of agrochemical and pharmacological activities. During our research on Actinomycetes from Brazilian mangroves, the ethyl acetate extract of Streptomyces sp. AMC 23 isolated from the red mangrove (Rhizophora mangle) rhizosphere produced a highly active compound against the microalga Chlorella vulgaris, often used to assess the phytotoxic activity. As a result, the bioassay-guided fractionation led to the isolation of the mixture of the known compounds bafilomycin B1 and bafilomycin B2. The chemical structures of bafilomycin B1 and bafilomycin B2 were established based on their spectroscopic data by infrared (IR), mass spectrometry (MS), (1)H nuclear magnetic resonance (NMR), gradient-enhanced heteronuclear multiple quantum coherence (gHMQC), and gradient-enhanced heteronuclear multiple-bond connectivity (gHMBC) as well as comparison with reference data from the literature. Moreover, it was also possible to identify other bafilomycins using non-chromatographic-dependent techniques (Tandem mass spectrometry). Additionally, this is the first report on the phytotoxic activity of bafilomycin B1.

New methymycin derivatives of Streptomyces venezuelae ATCC 15439 and their inhibitory effects on human T cell proliferation mediated by PMA/ionomycin.

Two new methymycin derivatives, 3'-demethylmethymycin (1) and 3'-demethyldeoxymethymycin (2), together with seven known ones (3-9), were obtained from the strain Streptomyces venezuelae ATCC 15439. Their structures were determined on the basis of IR, MS, 1D and 2D NMR data. In addition, the inhibitory effects of all the compounds on human T cell proliferation mediated by PMA/ionomycin were evaluated. The data suggested for the first time that methymycin derivatives have potential anti-inflammatory activity.

Biosynthesis of glycosylated derivatives of tylosin in Streptomyces venezuelae.

Streptomyces venezuelae YJ028, bearing a deletion of the entire biosynthetic gene cluster encoding the pikromycin polyketide synthases and desosamine biosynthetic enzymes, was used as a bioconversion system for combinatorial biosynthesis of glycosylated derivatives of tylosin. Two engineered deoxysugar biosynthetic pathways for the biosynthesis of TDP-3-O-demethyl-D-chalcose or TDP-Lrhamnose in conjunction with the glycosyltransferaseauxiliary protein pair DesVII/DesVIII were expressed in a S. venezuelae YJ028 mutant strain. Supplementation of each mutant strain capable of producing TDP-3-O-demethyl- D-chalcose or TDP-L-rhamnose with tylosin aglycone tylactone resulted in the production of the 3-O-demethyl- D-chalcose, D-quinovose, or L-rhamnose-glycosylated tylactone.

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

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.

Design and synthesis of pathway genes for polyketide biosynthesis.

In this chapter we describe novel methods for the design and assembly of synthetic pathways for the synthesis of polyketides and tailoring sugars. First, a generic design for type I polyketide synthase genes is presented that allows their facile assembly for the expression of chimeric enzymes in an engineered Escherichia coli host. The sequences of the synthetic genes are based on naturally occurring polyketide synthase genes but they are redesigned by custom-made software to optimize codon usage to maximize expression in E. coli and to provide a standard set of restriction sites to allow combinatorial assembly into unnatural enzymes. The methodology has been validated by building a large number of bimodular mini-PKSs that make easily assayed triketide products. Learning from the successful bimodules, a conceptual advance was made by assembling genes encoding functional trimodular enzymes, capable of making tetraketide products. Second, methods for the rapid assembly and exchange of sugar pathway genes into functional operons are described. The approach was validated by the assembly of the 15 genes for the synthesis of mycarose and desosamine in two operons, which yielded erythromycin C when coexpressed with the corresponding PKS genes. These methods are important enabling steps toward the goals of making designer drugs by polyketide synthase and sugar pathway engineering and, in the shorter term, producing by fermentation advanced intermediates for the synthesis of compounds that otherwise require large numbers of chemical steps.

Heterologous production of polyketides in bacteria.

Polyketide natural products are among the most important microbial metabolites in human medicine and are widely used to treat both acute and degenerative diseases. The need to develop new drugs has prompted the idea of using heterologous systems for the expression of polyketide biosynthetic pathways. The basic idea behind this approach is to use heterologous bacterial systems with better growth and genetic characteristics that could support better production of a certain compound than the original host or that could allow the generation of novel analogues through combinatorial biosynthesis. Moreover, these hosts could be used to express "cryptic" secondary metabolic pathways or serve as surrogate hosts in metagenomics experiments in order to find potential new bioactive compounds. In this chapter we discuss recent advances in the heterologous production of polyketides in bacteria and describe some methodological improvements of the systems.