Mycobacterial MMAR_2193 catalyzes O-methylation of diverse polyketide cores

O-methylation of small molecules is a common modification widely present in most organisms. Type III polyketides undergo O-methylation at hydroxyl end to play a wide spectrum of roles in bacteria, plants, algae, and fungi. Mycobacterium marinum harbours a distinctive genomic cluster with a type III pks gene and genes for several polyketide modifiers including a methyltransferase gene, mmar_2193. This study reports functional analyses of MMAR_2193 and reveals multi-methylating potential of the protein. Comparative sequence analyses revealed conservation of catalytically important motifs in MMAR_2193 protein. Homology-based structure-function and molecular docking studies suggested type III polyketide cores as possible substrates for MMAR_2193 catalysis. In vitro enzymatic characterization revealed the capability of MMAR_2193 protein to utilize diverse polyphenolic substrates to methylate several hydroxyl positions on a single substrate molecule. High-resolution mass spectrometric analyses identified multi-methylations of type III polyketides in cell-free reconstitution assays. Notably, our metabolomics analyses identified some of these methylated molecules in biofilms of wild type Mycobacterium marinum. This study characterizes a novel mycobacterial O-methyltransferase protein with multi-methylating enzymatic ability that could be exploited to generate a palette of structurally distinct bioactive molecules.

Genome-Wide Identification and Characterization of Caffeic Acid O-Methyltransferase Gene Family in Soybean

Soybean is one of the most important legumes, providing high-quality protein for humans. The caffeic acid O-methyltransferase (COMT) gene has previously been demonstrated to be a critical gene that regulates lignin production in plant cell walls and plays an important function in plant growth and development. However, the COMT gene family has not been studied in soybeans. In this study, 55 COMT family genes in soybean were identified by phylogenetic analysis and divided into two groups, I and II. The analysis of conserved domains showed that all GmCOMTs genes contained Methyltransferase-2 domains. Further prediction of cis-acting elements showed that GmCOMTs genes were associated with growth, light, stress, and hormonal responses. Eventually, based on the genomic data of soybean under different stresses, the results showed that the expression of GmCOMTs genes was different under different stresses, such as salt and drought stress. This study has identified and characterized the COMT gene family in soybean, which provides an important theoretical basis for further research on the biological functions of COMT genes and promotes revealing the role of GmCOMTs genes under stress resistance.

Influence of Catechol-O-Methyltransferase Gene Polymorphism on the Correlation between Alexithymia and Hypervigilance to Pain

The psychological characteristic of having difficulty expressing emotions, known as alexithymia, is associated with hypervigilance to pain and is considered one of the risk factors for chronic pain. The correlation between alexithymia and hypervigilance to pain can be observed even in healthy individuals. However, the factors influencing this correlation remain unknown. We explored the dopamine system, which is known to be involved in emotion and pain. The dopamine-degrading enzyme catechol-O-methyltransferase (COMT) has a genetic polymorphism known to influence dopamine metabolism in the prefrontal cortex. COMT polymorphism reportedly affects various aspects of pain and increases pain sensitivity in Met allele carriers. Therefore, we investigated whether the correlation between alexithymia and hypervigilance to pain is influenced by COMT polymorphism in healthy individuals. The results revealed a significant positive correlation between the “difficulty describing feelings” of the 20-item Toronto Alexithymia Scale and the “attention to changes in pain” of the pain vigilance and awareness questionnaire in COMT Met carriers but not in Val/Val individuals. This finding suggests that the correlation between alexithymia and hypervigilance to pain is influenced by COMT polymorphism.

Evaluation of the Association of COMT Rs4680 Polymorphism with Swimmers’ Competitive Performance

Swimmers’ competitive performance is a result of complicated interactions between physiological, biochemical, physical and psychological factors, all of which are strongly affected by water. Recently, great attention has been paid to the role of genetic factors such as the catechol-O-methyltransferase gene (COMT) influencing motivation, emotions, stress tolerance, self-control, sleep regulation, pain processing and perception, addictive behaviour and neurodegeneration, which may underlie differences in achieving remarkable results in sports competition. Thus, this study was performed to investigate the association between the COMT Val158Met (rs4680) polymorphism and athletic performance in Caucasian swimmers. A total of 225 swimmers (171 short distance (SDS) and 54 long distance swimmers (LDS)) of national or international competitive standard and 379 unrelated sedentary controls were genotyped using real-time polymerase chain reaction (real-time PCR). We found no significant differences in genotypic or allelic distributions between (1) male and female athletes; (2) SDS and LDS; (3) all athletes and sedentary controls (under codominant, dominant, recessive, and overdominant genetic models). No association was found between the COMT rs4680 polymorphism and elite swimming athlete status of the studied population. However, more replication studies are needed.

In vivo emergence of high-level resistance during treatment reveals the first identified mechanism of amphotericin B resistance in Candida auris

Candida auris has emerged as a healthcare-associated and multidrug-resistant fungal pathogen of great clinical concern. While as much as 50% of C. auris clinical isolates are reported to be resistant to amphotericin B, to date, no mechanisms contributing to this resistance have been identified. We report here mutations in the C. auris sterol-methyltransferase gene, ERG6, as the first identified mechanism of amphotericin B resistance in this emerging pathogen and describe the clinical case in which this high-level amphotericin B resistance was acquired in vivo during therapy. Whole genome sequencing revealed the four C. auris isolates obtained from this single patient case to be genetically related and identified a mutation in ERG6 as being associated with amphotericin B resistance. Cas9-mediated genetic manipulations confirmed this mutation alone to confer a >32-fold increase in amphotericin B resistance, and comprehensive sterol profiling revealed an abrogation of ergosterol biosynthesis and a corresponding accumulation of cholesta-type sterols in isolates and strains harboring the clinically derived ERG6 mutation. Together these findings represent the first significant step forward in the understanding of clinical amphotericin B resistance in C. auris.

Characterization of Methyltransferase from Apramycin Biosynthetic Gene Cluster and Improvement Production of Apramycin in Engineered Streptoalloteichus Tenebrarius

BackgroundApramycin is a structurally unique aminoglycoside, used in veterinary medicine or the treatment of Salmonella, Escherichia coli and Pasteurella multocida infections in farm. Although discovered and used many years ago, many biosynthetic steps of apramycin are still obscure. ResultsIn this study, we identified a HemK family methyltransferase, aprI, involved in apramycin biosynthesis. The function of aprI was studied by using gene disruption and biochemical experiments, and a new aminoglycoside antibiotic demethyl-apramycin was purified from aprI disruption strain. Experiments proved that AprI converted demethyl-aprosamine to aprosamine in vitro. Based on this, the apramycin production strain was improved by overexpression the AprI to decrease the impurity production. ConclusionsWe have identified aprI is a 7’-N-methyltransferase gene in apramycin biosynthesis and confirmed the substrate of methyltransferase. Engineering of aprI resulted in a strain producing a new aminoglycoside demethyl-apramycin and apramycin mono-producing strain with less impurity production. Finally, the yield of demethyl-apramycin in apramycin mono-producing strain decreased from 196±36 mg/L to 51±9 mg/L, and the yield of apramycin increased from 2227±320 mg/L to 2331±210 mg/L.