Open AccessHeterologous Expression of Cryptomaldamide in a Cyanobacterial Host
Author(s) -
Arnaud Taton,
Andrew K. Ecker,
Brienna Diaz,
Nathan A. Moss,
Brooke Anderson,
Raphael Reher,
Tiago Leão,
Ryan Simkovsky,
Pieter C. Dorrestein,
Lena Gerwick,
William H. Gerwick,
James W. Golden
Publication year - 2020
Publication title -
acs synthetic biology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 2.156
H-Index - 66
ISSN - 2161-5063
DOI - 10.1021/acssynbio.0c00431
Subject(s) - biology , nonribosomal peptide , gene cluster , heterologous expression , cyanobacteria , cylindrospermopsis raciborskii , gene , anabaena , nostoc , heterologous , synthetic biology , genetics , computational biology , biosynthesis , recombinant dna , bacteria
Filamentous marine cyanobacteria make a variety of bioactive molecules that are produced by polyketide synthases, nonribosomal peptide synthetases, and hybrid pathways that are encoded by large biosynthetic gene clusters. These cyanobacterial natural products represent potential drug leads; however, thorough pharmacological investigations have been impeded by the limited quantity of compound that is typically available from the native organisms. Additionally, investigations of the biosynthetic gene clusters and enzymatic pathways have been difficult due to the inability to conduct genetic manipulations in the native producers. Here we report a set of genetic tools for the heterologous expression of biosynthetic gene clusters in the cyanobacteria Synechococcus elongatus PCC 7942 and Anabaena ( Nostoc ) PCC 7120. To facilitate the transfer of gene clusters in both strains, we engineered a strain of Anabaena that contains S. elongatus homologous sequences for chromosomal recombination at a neutral site and devised a CRISPR-based strategy to efficiently obtain segregated double recombinant clones of Anabaena . These genetic tools were used to express the large 28.7 kb cryptomaldamide biosynthetic gene cluster from the marine cyanobacterium Moorena ( Moorea ) producens JHB in both model strains. S. elongatus did not produce cryptomaldamide; however, high-titer production of cryptomaldamide was obtained in Anabaena . The methods developed in this study will facilitate the heterologous expression of biosynthetic gene clusters isolated from marine cyanobacteria and complex metagenomic samples.