Final Report Summary - LYNGBYA-KENYA (Cyp-450 biosynthesis of Lyngbya majuscula natural products)
EXECUTIVE SUMMARY
A sustainable supply of marine natural products for potential therapeutics is one of the greatest challenges facing drug discovery efforts today, especially during clinical trials. Nearly 300 compounds with therapeutic potential have been isolated from the tropical marine cyanobacterium Lyngbya majuscula. However there are considerable concerns regarding the real source of this large number of natural products attributed to L. majuscula. This project focused on the cytochrome p450 biosynthesis of L. majuscula natural products namely the modular cyclodepsipeptides homodolastatin 16 (HMDS 16) and antanapeptin A (ANTAP A). L. majuscula was collected from Shimoni, Kenya in April 2012. The aim was to identify if these compounds originate from the cyanobacteria or bacteria cohabiting with it or both. Bacteria representing γ-proteobacteria, Firmicutes, and α–proteobacteria were isolated from the cyanobacteria. Non-ribosomal peptide synthetase (NRPS) and polyketide synthase (PKS) screens of bacteria with known complete genome sequences using bioinformatics tools showed in this study that the modular assembly lines for these bacteria are inconsistent with those of HMDS 16 and ANTAP A. However, Bacillus licheniformis and Marinobacterium stanieri synthesise the β amino acid dolamethleuline and Klebsiella oxytoca is involved with the biosynthesis of the unusual dolaphenvaline amino acid in HMDS 16 putatively. Profiling for the cyclodepsipeptides in bacteria supernatants using liquid chromatograph mass spectrometry (LCMS) confirmed the presence of HMDS 16, its analogue dolastatin 16, and ANTAP A in L. majuscula but not in the bacteria. Subsequently such leads aided the prospects for obtaining the complete genome sequence of L. majuscula from its metagenome and in identifying gene clusters encoding for HMDS 16 and ANTAP A. The cloning and heterologous expression of the gene clusters for these especially important anticancer agents is the goal that we aimed to attain by the end of the program.
A sustainable supply of marine natural products for potential therapeutics is one of the greatest challenges facing drug discovery efforts today, especially during clinical trials. Nearly 300 compounds with therapeutic potential have been isolated from the tropical marine cyanobacterium Lyngbya majuscula. However there are considerable concerns regarding the real source of this large number of natural products attributed to L. majuscula. This project focused on the cytochrome p450 biosynthesis of L. majuscula natural products namely the modular cyclodepsipeptides homodolastatin 16 (HMDS 16) and antanapeptin A (ANTAP A). L. majuscula was collected from Shimoni, Kenya in April 2012. The aim was to identify if these compounds originate from the cyanobacteria or bacteria cohabiting with it or both. Bacteria representing γ-proteobacteria, Firmicutes, and α–proteobacteria were isolated from the cyanobacteria. Non-ribosomal peptide synthetase (NRPS) and polyketide synthase (PKS) screens of bacteria with known complete genome sequences using bioinformatics tools showed in this study that the modular assembly lines for these bacteria are inconsistent with those of HMDS 16 and ANTAP A. However, Bacillus licheniformis and Marinobacterium stanieri synthesise the β amino acid dolamethleuline and Klebsiella oxytoca is involved with the biosynthesis of the unusual dolaphenvaline amino acid in HMDS 16 putatively. Profiling for the cyclodepsipeptides in bacteria supernatants using liquid chromatograph mass spectrometry (LCMS) confirmed the presence of HMDS 16, its analogue dolastatin 16, and ANTAP A in L. majuscula but not in the bacteria. Subsequently such leads aided the prospects for obtaining the complete genome sequence of L. majuscula from its metagenome and in identifying gene clusters encoding for HMDS 16 and ANTAP A. The cloning and heterologous expression of the gene clusters for these especially important anticancer agents is the goal that we aimed to attain by the end of the program.