Project description:Several soil organisms are known to be capable of growth on caprolactam as soil carbon and nitrogen source, but the enzymes of the catabolic pathway have not been described. We isolated a caprolactam-degrading strain of Pseudomonas jessenni, and identified genes putatively involved in the caprolactam metabolism using quantitative mass spectrometry-based proteomics. This led to the discovery of a caprolactamase and an aminotransferase that are involved in the initial steps of caprolactam conversion. Additionally, various proteins were identified likely to be involved in later steps of the pathway. The identified caprolactamase consisted of 2 subunits and demonstrated high sequence identity to the 5-oxoprolinases. E. coli expressing this caprolactamase did not convert 5-oxoproline but was able to hydrolyze caprolactam to form 6-aminohexanoic acid in an ATP dependent manner. Characterization of the aminotransferase revealed that the enzyme deaminated 6-aminohexanoic acid to produce 6-oxohexanoate with pyruvate as amino acceptor. The amino acid sequence of the aminotransferase demonstrated high similarity to subgroup II ω-aminotransferases of the PLP fold type I proteins. Finally, analyses of the genome sequence demonstrated the presence of a caprolactam catabolism gene cluster consisting of all genes involved in the conversion of caprolactam to adipate.
Project description:Pseudomonas aeruginosa is an opportunistic pathogen which causes acute and chronic infections that are difficult to treat. Comparative genomic analysis has showed a great genome diversity among P. aeruginosa clinical strains and revealed important regulatory traits during chronic adaptation. While current investigation of epigenetics of P. aeruginosa is still lacking, understanding the epigenetic regulation may provide biomarkers for diagnosis and reveal important regulatory mechanisms. The present study focused on characterization of DNA methyltransferases (MTases) in a chronically adapted P. aeruginosa clinical strain TBCF10839. Single-molecule real-time sequencing (SMRT-seq) was used to characterize the methylome of TBCF. RCCANNNNNNNTGAR and TRGANNNNNNTGC were identified as target motifs of DNA MTases, M.PaeTBCFI and M.PaeTBCFII, respectively.
Project description:Indole-3-acetic acid (IAA), knows as common plant hormone, is one of the most distributed indole derivatives in the environment. A novel strain, which was able to use IAA as sole source of carbon and nitrogen, was isolated from farm soil, identified and classified as Pseudomonas composti LY1 based on 16S rRNA sequence and genome analysis. The optimal growth conditions for LY1 with IAA are characterized. Proteome profile of strain LY1 to IAA and citrate were analyzed and compared using label free strategy with LC-MS/MS.
