Synthesis and Evaluation of Acridone and Xanthone Epoxides with Anti-MRSA and Anti-MSSA Activity

A series of acridone and xanthone-based compounds bearing 1,2-epoxypropyl or 1,2-propanediol substituents were synthesized and evaluated for activity against MRSA and MSSA bacterial strains. The results indicate a correlation exists between the number of epoxide groups and activity, with peak MIC values observed for bis-epoxy derivatives. Both activity and heathy cell toxicity was shown to decrease with the addition of a third epoxy group. The corresponding ring-opened diol analogs were devoid of activity, demonstrating the critical function of the epoxide in mediating antimicrobial activity. The most active compounds were also screened using a regulated antisense RNA expression library. The results show no increase in activity against cells sensitized by down-regulation of the most common drug targets, including DNA gyrase, DNA topoisomerase, tRNA synthetase, and the fatty acid biosynthesis pathway. The compounds are postulated to function as membrane disrupting agents, similar to the xanthone natural product α-mangostin.

Genome Sequencing and Analysis of Thraustochytriidae sp. SZU445 Provides Novel Insights into the Polyunsaturated Fatty Acid Biosynthesis Pathway

Thraustochytriidae sp. have broadly gained attention as a prospective resource for the production of omega-3 fatty acids production in significant quantities. In this study, the whole genome of Thraustochytriidae sp. SZU445, which produces high levels of docosapentaenoic acid (DPA) and docosahexaenoic acid (DHA), was sequenced and subjected to protein annotation. The obtained clean reads (63.55 Mb in total) were assembled into 54 contigs and 25 scaffolds, with maximum and minimum lengths of 400 and 0.0054 Mb, respectively. A total of 3513 genes (24.84%) were identified, which could be classified into six pathways and 44 pathway groups, of which 68 genes (1.93%) were involved in lipid metabolism. In the Gene Ontology database, 22,436 genes were annotated as cellular component (8579 genes, 38.24%), molecular function (5236 genes, 23.34%), and biological process (8621 genes, 38.42%). Four enzymes corresponding to the classic fatty acid synthase (FAS) pathway and three enzymes corresponding to the classic polyketide synthase (PKS) pathway were identified in Thraustochytriidae sp. SZU445. Although PKS pathway-associated dehydratase and isomerase enzymes were not detected in Thraustochytriidae sp. SZU445, a putative DHA- and DPA-specific fatty acid pathway was identified.

Evaluating the effects of synthetic POM cycles and NAD kinase expression on fatty alcohol production in Saccharomyces cerevisiae

Background: NADPH-dependent enzymes play important roles in many anabolic reactions and the availability of redox cofactors can influence metabolic flux ultimately influencing titers of bioproducts produced by engineered microbial cells. This may be especially true of oleochemical production when carbon flux through the highly NADPH-dependent fatty acid biosynthesis pathway is increased. While pathway specific approaches are often applied to counter redox imbalance, a study evaluating generalized approaches to improved NADPH availability is lacking in Saccharomyces cerevisiae . Results: Here, we have created four unique synthetic Pyruvate-Oxaloacetate-Malate “POM” cycles consisting of either of the endogenous isoforms of pyruvate carboxylase ( PYC1 or PYC2 ), a modified version of malate dehydrogenase ( ‘MDH1 or ‘MDH2 ), and a truncated cytosolic form of the endogenous malic enzyme ( sMAE1 ). Only the POM cycle that combined expression of PYC1 , ‘MDH2 , and sMAE1 increased the titer of fatty alcohols produced; however, it did so in two unique fatty alcohol producing strains. In a FAS1 overexpression background, expression of this synthetic POM cycle increased fatty alcohol titers by 40% from 49.0 ± 2.2 mg/L to 68.6 ± 3.3 mg/L and showed similar results in a zwf1 deletion strain. The effect of overexpression of the endogenous NAD+ kinases UTR1 , YEF1 , and a cytosolic version of POS5 were also tested. We found that expression of POS5c resulted in an ~35% increase in fatty alcohol titer, while the overexpression of the UTR1 or YEF1 did not significantly influence titers. In these minimally engineered cells, combined overexpression of PYC1 , ‘ MDH2 , sMAE1 and POS5c did not further increase titers Conclusions: Overexpression of PYC1 in conjunction with ‘MDH2 and sMAE1 results in a synthetic POM cycle which can be utilized to improve fatty alcohol production in engineered strains of S. cerevisiae . Additionally, overexpression of a truncated version of POS5 ( POS5c ) results in similar increases in fatty alcohol production. These findings may serve to provide a generalized mechanism to increase NADPH production in engineered cells, resulting in increased bioproduct titers.

Production of Bioactive Compounds by Food Associated Galactomyces geotrichum 38, as Determined by Proteome Analysis

Fried cottage cheese is a dairy product, popular in some parts of Poland. Proteomic analysis of a culture of the mold Galactomyces geotrichum 38 isolated from fried cottage cheese was performed using UHPLC/MS. From the proteins identified, we selected those involved in the biosynthesis of bioactive compounds and those useful in industry. In the G. geotrichum 38 culture, the production quantities of vitamin B2 (224 μg/L), ergosterol (54.63 mg/kg), and trehalose (0.91 g/L) were determined by HPLC. The identified proteins were also used to prepare a hypothetical fatty acid biosynthesis pathway, and the percentage of individual sphingolipids in the culture was determined. Sphingolipids are also bioactive compounds. During culturing of G. geotrichum 38, the percentage of three sphingolipids increased. The last step of the research was to prepare a model of fried cottage cheese. The mold G. geotrichum 38, used in the process of ripening fried cottage cheese, synthesized vitamin B2 and erogsterol, which influenced the nutritional value of the product.

Genome-wide characterization of Phytophthora infestans metabolism: a systems biology approach

Genome-scale metabolic models (GEMs) provide a functional view of the complex network of biochemical reactions in the living cell. Initially mainly applied to reconstruct the metabolism of model organisms, the availability of increasingly sophisticated reconstruction methods and more extensive biochemical databases now make it possible to reconstruct GEMs for less characterized organisms as well, and have the potential to unravel the metabolism in pathogen-host systems. Here we present a GEM for the oomycete plant pathogen Phytophthora infestans as a first step towards an integrative model with its host. We predict the biochemical reactions in different cellular compartments and investigate the gene-protein-reaction associations in this model to get an impression of the biochemical capabilities of P. infestans. Furthermore, we generate life stage-specific models to place the transcriptomic changes of genes encoding metabolic enzymes into a functional context. In sporangia and zoospores there is an overall downregulation, most strikingly reflected in the fatty acid biosynthesis pathway. To investigate the robustness of the GEM, we simulate gene deletions to predict which enzymes are essential for in vitro growth. While there is room for improvement, this first model is an essential step towards an understanding of P. infestans and its interactions with plants as a system, which will help to formulate new hypotheses on infection mechanisms and disease prevention.