is a large multifunctional enzyme that has modular s


is a large multifunctional enzyme that has modular buy FG-4592 structures [4]. Each NRPS module catalyses the incorporation of a specific substrate into the growing product. A typical module consists of three enzymatic domains, namely, adenylation (A), thiolation (T; also known as peptidyl carrier protein), and condensation (C) domains. The A domain selects and activates a specific amino acid substrate, the T domain is responsible for tethering the activated substrate to the 4′-phosphopanthetheinyl cofactor, and the C domain catalyses peptide bond formation between the elongating peptide and a new amino acid. In addition to these core domains, the terminal thioesterase (TE) and epimerisation (E) domains, as well as several other tailoring domains, may also be present in NRPS modules. The order of modules of an NRPS is, in many cases, collinear to the amino acid sequence of the corresponding peptide product. The collinearity rule Vorinostat mouse of NRPS systems combined with knowledge of the specificity-conferring code of A domain allow for the prediction and amino acid modification of peptide fragments synthesised by corresponding NRPS

[5]. However, few NRPS sequences have been extensively described in comparison with the number of known peptide products, limiting the study of the principles of non-ribosomal peptide synthesis and the development of new bioactive peptides by genetic engineering. In this study, we identified PRKACG and analysed a gene cluster involved in

the biosynthesis of pelgipeptin and provided biochemical data for the collinearity of this peptide assembly line. Methods Bacteria learn more strains and culture conditions P. elgii B69, isolated from a soil sample [1], was cultured in nutrient broth. E. coli DH5α, for gene manipulation, and E. coli BL21 (DE3), for overexpression of recombinant proteins, were cultivated on Luria-Bertani medium. Identification and in silico analysis of plp gene cluster in P. elgii B69 The draft genome sequence of the strain was used to build a database in Bioedit to identify the putative NRPS genes in P. elgii B69 (http://​www.​mbio.​ncsu.​edu/​BioEdit/​bioedit.​html). The first and second C domains of PmxE (GenBank EU371992), which is a polymyxin synthetase subunit, were compared with the created database using local BLAST searches [6] as implemented in Bioedit. Amino acid sequence homology searches were performed using the BLAST server at the National Centre for Biotechnology Information (NCBI, http://​www.​ncbi.​nlm.​nih.​gov/​BLAST/​) site. NRPS domains were identified by PKS/NRPS analysis (http://​nrps.​igs.​umaryland.​edu/​nrps/​) [7]. Prediction of 10 amino acids located at the substrate-binding pocket of the A domain and substrate specificity prediction were performed using the web-based program NRPS predictor (http://​ab.​inf.​uni-tuebingen.​de/​software/​NRPSpredictor/​) [8].

Comments are closed.