Steroidi i poliketidi iz kore biljke Uvaria hamiltonii

Two known steroids, stigmasterol and 6beta-hydroxystigmasta-4,22-dien-3-one (1) and two unusual polyketides, cis-4-hydroxymellein (2) and trans-4-hydroxymellein (3) were isolated from the stem bark of Uvaria hamiltonii. The structures of the compounds were elucidated independently by high-resolution 2D-NMR techniques and confirmed by comparison with previously reported values.Dva poznata steroida, stigmasterol i 6-hidroksistigmasta-4,22-dien-3-on (1) i dva neuobičajena poliketida, cis-4-hidroksimelein (2) i trans-4-hidroksimelein (3) izolirana su iz kore biljke Uvaria hamiltonii. Strukture spojeva određene su neovisno 2D-NMR spektroskopijom visoke rezolucije i usporedbom s literaturnim podacima

Natural product diversity of actinobacteria in the Atacama Desert

Natural Product diversityPeer reviewedPublisher PD

Aspergillus westerdijkiae polyketide synthase gene “aoks1” is involved in the biosynthesis of ochratoxin A

OchratoxinA (OTA) is a potential nephrotoxic, teratogenic, immunogenic, hepatotoxic and carcinogenic mycotoxin, produced by Aspergillus westerdijkiae NRRL 3174. Herein we describe the characterization of a putative OTA-polyketide synthasegene “aoks1”, cloned by using gene walking approach. The predicted amino acid sequence of the 2 kb clone display 34–60% similarities to different polyketide synthasegenes including lovastatine biosynthesis gene “lovb” in A. terreus, compactin biosynthesis gene “mlcA” in Penicillium citrinum and OTA biosynthesis gene “otapksPN” in P. nordicum. Based on the reverse transcription PCR and kinetic secondary metabolites production studies, aoks1 expression was found to be associated with OTA biosynthesis. Further a mutant, in which the aoks1gene was inactivated by Escherichia coli hygromycin B phosphotransferase gene, lost the capacity to produce OTA, but still producing mellein. To our knowledge this report describes for the first time characterization of a gene involved in OTA biosynthesis, with the information about mellein which was proposed in the literature to be an intermediate OTA. This study also suggests that aoks1 may be the second polyketide synthase gene required for OTA biosynthesis in A. westerdijkiae NRRL 3174

Biocatalytic Synthesis of Stereospecific Triketide Lactones using Polyketide Synthases

Polyketide synthases are modular enzymes that create and modify large acyl chains. The domains and modules of polyketide synthases allow us to create molecules that resemble naturally occurring products by applying a biocatalytic in vitro in vivo approach to a diketide acyl chain. We showed that a triketide lactone of desired stereochemistry could be made using a domain and module from the polyketide synthase found in Saccharopolyspora erythraea, 6-Deoxyerythronolide B Synthase. Future projects will explore this approach using different domains and modules.Biochemistr

Isolation of pigment cell specific genes in the sea urchin embryo by differential macroarray screening

New secondary mesenchyme specific genes, expressed exclusively in pigment cells, were isolated from sea urchin embryos using a differential screening of a macroarray cDNA library. The comparison was performed between mRNA populations of embryos having an expansion of the endo-mesodermal territory and embryos blocked in secondary mesenchyme specification. To be able to isolate transcripts with a prevalence down to five copies per cell, a subtractive hybridization procedure was employed. About 400 putative positive clones were identified and sequenced from the 5' end. Gene expression analysis was carried out on a subset of 66 clones with real time quantitative PCR and 40 clones were positive. This group of clones contained sequences highly similar to: the transcription factor glial cells missing (gcm); the polyketide synthase gene cluster (pks-gc); three different members of the flavin-containing monooxygenase gene family (fmo); and a sulfotransferase gene (sult). Using whole mount in situ hybridization, it was shown that these genes are specifically expressed in pigment cells. A functional analysis of the S. purpuratus pks and of one S. purpuratus fmo was carried out using antisense technology and it was shown that their expression is necessary for the biosynthesis of the sea urchin pigment echinochrome. The results suggest that S. purpuratus pks, fmo and sult could belong to a differentiation gene battery of pigment cells

MultiMetEval: comparative and multi-objective analysis of genome-scale metabolic models

Comparative metabolic modelling is emerging as a novel field, supported by the development of reliable and standardized approaches for constructing genome-scale metabolic models in high throughput. New software solutions are needed to allow efficient comparative analysis of multiple models in the context of multiple cellular objectives. Here, we present the user-friendly software framework Multi-Metabolic Evaluator (MultiMetEval), built upon SurreyFBA, which allows the user to compose collections of metabolic models that together can be subjected to flux balance analysis. Additionally, MultiMetEval implements functionalities for multi-objective analysis by calculating the Pareto front between two cellular objectives. Using a previously generated dataset of 38 actinobacterial genome-scale metabolic models, we show how these approaches can lead to exciting novel insights. Firstly, after incorporating several pathways for the biosynthesis of natural products into each of these models, comparative flux balance analysis predicted that species like Streptomyces that harbour the highest diversity of secondary metabolite biosynthetic gene clusters in their genomes do not necessarily have the metabolic network topology most suitable for compound overproduction. Secondly, multi-objective analysis of biomass production and natural product biosynthesis in these actinobacteria shows that the well-studied occurrence of discrete metabolic switches during the change of cellular objectives is inherent to their metabolic network architecture. Comparative and multi-objective modelling can lead to insights that could not be obtained by normal flux balance analyses. MultiMetEval provides a powerful platform that makes these analyses straightforward for biologists. Sources and binaries of MultiMetEval are freely available from https://github.com/PiotrZakrzewski/MetEv​al/downloads

Structure determination of a 4-pyrone from the liverwort <i>Plagiochila bifaria</i> (Sw.) Lindenb. (Plagiochilaceae)

The centenary of the presentation in 1907 of the "polyketide" hypothesis by Collie, along with his use of a 4-pyrone as an example, is marked by reporting the discovery and isolation of 2-(3,4-dimethoxyphenethyl)-6-methyl-4-pyrone from &lt;i&gt;Plagiochila bifaria&lt;/i&gt;. This compound dominates the phytochemical profile of the deuteriochloroform extract of a Venezuelan specimen of &lt;i&gt;P. bifaria&lt;/i&gt; and the structure was determined using spectroscopic techniques, especially 2D NMR. This natural product is novel because it contains what appears to be the first example of a monocyclic 4-pyrone that has a polyketide origin. The hypothetical pathway for cyclization of a triketocarboxylic acid to a 4-pyrone was known to be applicable to model systems but no examples of natural products from this route were known. This compound is the same as one of a series of compounds observed previously in an extract of a Brazilian specimen of &lt;i&gt;P. bifaria&lt;/i&gt;; the working structures that were proposed earlier require revision. The mass spectral characteristics are the same as those reported (parent and base peak) in 1987 for the major component observed in the GC-MS profile of an extract of a specimen of &lt;i&gt;P. bifaria&lt;/i&gt; from Peru. The roles played by Birch and Robinson in the renaissance of the polyketide hypothesis almost fifty years after its initial launch are considered. Based on evidence from their publications, they worked independently of each other. It appears Robinson always had knowledge of Collie's hypothesis when developing ideas about structural relations of natural products whereas Birch initially was unaware of both Collie's and Robinson's ideas on the subject

Simultaneous Effect of Temperature and Irradiance on Growth and Okadaic Acid Production from the Marine Dinoflagellate Prorocentrum belizeanum

Benthic marine dioflagellate microalgae belonging to the genus Prorocentrum are a major source of okadaic acid (OA), OA analogues and polyketides. However, dinoflagellates produce these valuable toxins and bioactives in tiny quantities, and they grow slowly compared to other commercially used microalgae. This hinders evaluation in possible large-scale applications. The careful selection of producer species is therefore crucial for success in a hypothetical scale-up of culture, as are appropriate environmental conditions for optimal growth. A clone of the marine toxic dinoflagellate P. belizeanum was studied in vitro to evaluate its capacities to grow and produce OA as an indicator of general polyketide toxin production under the simultaneous influence of temperature (T) and irradiance (I0). Three temperatures and four irradiance levels were tested (18, 25 and 28 °C; 20, 40, 80 and 120 µE·m−2·s−1), and the response variables measured were concentration of cells, maximum photochemical yield of photosystem II (PSII), pigments and OA. Experiments were conducted in T-flasks, since their parallelepipedal geometry proved ideal to ensure optically thin cultures, which are essential for reliable modeling of growth-irradiance curves. The net maximum specific growth rate (µm) was 0.204 day−1 at 25 °C and 40 µE·m−2·s−1. Photo-inhibition was observed at I0 > 40 μEm−2s−1, leading to culture death at 120 µE·m−2·s−1 and 28 °C. Cells at I0 ≥ 80 µE·m−2·s−1 were photoinhibited irrespective of the temperature assayed. A mechanistic model for µm-I0 curves and another empirical model for relating µm-T satisfactorily interpreted the growth kinetics obtained. ANOVA for responses of PSII maximum photochemical yield and pigment profile has demonstrated that P. belizeanum is extremely light sensitive. The pool of photoprotective pigments (diadinoxanthin and dinoxanthin) and peridinin was not able to regulate the excessive light-absorption at high I0-T. OA synthesis in cells was decoupled from optimal growth conditions, as OA overproduction was observed at high temperatures and when both temperature and irradiance were low. T-flask culture observations were consistent with preliminary assays outdoors