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Development of a high-throughput strategy for discovery of potent analogues of antibiotic lysocin E.

ABSTRACT: Lysocin E, a 37-membered natural depsipeptide, induces rapid bacteriolysis in methicillin-resistant Staphylococcus aureus via a unique menaquinone-dependent mechanism, presenting a promising therapeutic lead. Despite the great medical importance, exploring the potential utility of its derivatives as new platform structures for antibiotic development has remained a significant challenge. Here, we report a high-throughput strategy that enabled the preparation of thousands of analogues of lysocin E and large-scale structure-activity relationship analyses. We integrate 26-step total synthesis of 2401 cyclic peptides, tandem mass spectrometry-sequencing, and two microscale activity assays to identify 23 candidate compounds. Re-synthesis of these candidates shows that 11 of them (A1-A11) exhibit antimicrobial activity superior or comparable to that of lysocin E, and that lysocin E and A1-A11 share L-Leu-6 and L-Ile-11. Therefore, the present strategy allows us to efficiently decipher biologically crucial residues and identify potentially useful agents for the treatment of infectious diseases.

SUBMITTER: Itoh H

PROVIDER: S-EPMC6611794 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:BackgroundSingle nucleotide polymorphisms (SNP) have been applied as important molecular markers in genetics and breeding studies. The rapid advance of next generation sequencing (NGS) provides a high-throughput means of SNP discovery. However, SNP development is limited by the availability of reliable SNP discovery methods. Especially, the optimum assembler and SNP caller for accurate SNP prediction from next generation sequencing data are not known.ResultsHerein we performed SNP prediction based on RNA-seq data of peach and mandarin peel tissue under a comprehensive comparison of two paired-end read lengths (125 bp and 150 bp), five assemblers (Trinity, IDBA, oases, SOAPdenovo, Trans-abyss) and two SNP callers (GATK and GBS). The predicted SNPs were compared with the authentic SNPs identified via PCR amplification followed by gene cloning and sequencing procedures. A total of 40 and 240 authentic SNPs were presented in five anthocyanin biosynthesis related genes in peach and in nine carotenogenic genes in mandarin. Putative SNPs predicted from the same RNA-seq data with different strategies led to quite divergent results. The rate of false positive SNPs was significantly lower when the paired-end read length was 150 bp compared with 125 bp. Trinity was superior to the other four assemblers and GATK was substantially superior to GBS due to a low rate of missing authentic SNPs. The combination of assembler Trinity, SNP caller GATK, and the paired-end read length 150 bp had the best performance in SNP discovery with 100% accuracy both in peach and in mandarin cases. This strategy was applied to the characterization of SNPs in peach and mandarin transcriptomes.ConclusionsThrough comparison of authentic SNPs obtained by PCR cloning strategy and putative SNPs predicted from different combinations of five assemblers, two SNP callers, and two paired-end read lengths, we provided a reliable and efficient strategy, Trinity-GATK with 150 bp paired-end read length, for SNP discovery from RNA-seq data. This strategy discovered SNP at 100% accuracy in peach and mandarin cases and might be applicable to a wide range of plants and other organisms.

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Development of a high-throughput strategy for discovery of potent analogues of antibiotic lysocin E.

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