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Bruns, H, Crusemann M, Letzel AC, Alanjary M, McInerney JO, Jensen PR, Schulz S, Moore BS, Ziemert N.  2018.  Function-related replacement of bacterial siderophore pathways. Isme Journal. 12:320-329.   10.1038/ismej.2017.137   AbstractWebsite

Bacterial genomes are rife with orphan biosynthetic gene clusters (BGCs) associated with secondary metabolism of unrealized natural product molecules. Often up to a tenth of the genome is predicted to code for the biosynthesis of diverse metabolites with mostly unknown structures and functions. This phenomenal diversity of BGCs coupled with their high rates of horizontal transfer raise questions about whether they are really active and beneficial, whether they are neutral and confer no advantage, or whether they are carried in genomes because they are parasitic or addictive. We previously reported that Salinispora bacteria broadly use the desferrioxamine family of siderophores for iron acquisition. Herein we describe a new and unrelated group of peptidic siderophores called salinichelins from a restricted number of Salinispora strains in which the desferrioxamine biosynthesis genes have been lost. We have reconstructed the evolutionary history of these two different siderophore families and show that the acquisition and retention of the new salinichelin siderophores co- occurs with the loss of the more ancient desferrioxamine pathway. This identical event occurred at least three times independently during the evolution of the genus. We surmise that certain BGCs may be extraneous because of their functional redundancy and demonstrate that the relative evolutionary pace of natural pathway replacement shows high selective pressure against retention of functionally superfluous gene clusters.

Letzel, AC, Li J, Amos GCA, Millan-Aguinaga N, Ginigini J, Abdelmohsen UR, Gaudencio SP, Ziemert N, Moore BS, Jensen PR.  2017.  Genomic insights into specialized metabolism in the marine actinomycete Salinispora. Environmental Microbiology. 19:3660-3673.   10.1111/1462-2920.13867   AbstractWebsite

Comparative genomics is providing new opportunities to address the diversity and distributions of genes encoding the biosynthesis of specialized metabolites. An analysis of 119 genome sequences representing three closely related species of the marine actinomycete genus Salinispora reveals extraordinary biosynthetic diversity in the form of 176 distinct biosynthetic gene clusters (BGCs) of which only 24 have been linked to their products. Remarkably, more than half of the BGCs were observed in only one or two strains, suggesting they were acquired relatively recently in the evolutionary history of the genus. These acquired gene clusters are concentrated in specific genomic islands, which represent hot spots for BGC acquisition. While most BGCs are stable in terms of their chromosomal position, others migrated to different locations or were exchanged with unrelated gene clusters suggesting a plug and play type model of evolution that provides a mechanism to test the relative fitness effects of specialized metabolites. Transcriptome analyses were used to address the relationships between BGC abundance, chromosomal position and product discovery. The results indicate that recently acquired BGCs can be functional and that complex evolutionary processes shape the micro-diversity of specialized metabolism observed in closely related environmental bacteria.

Crusemann, M, O'Neill EC, Larson CB, Melnik AV, Floros DJ, da Silva RR, Jensen PR, Dorrestein PC, Moore BS.  2017.  Prioritizing natural product diversity in a collection of 146 bacterial strains based on growth and extraction protocols. Journal of Natural Products. 80:588-597.   10.1021/acsjnatprod.6b00722   AbstractWebsite

In order to expedite the rapid and efficient discovery and isolation of novel specialized metabolites, while minimizing the waste of resources on rediscovery of known compounds, it is crucial to develop efficient approaches for strain prioritization, rapid dereplication, and the assessment of favored cultivation and extraction conditions. Herein we interrogated bacterial strains by systematically evaluating cultivation and extraction parameters with LC-MS/MS analysis and subsequent dereplication through the Global Natural Product Social Molecular Networking (GNPS) platform. The developed method is fast, requiring minimal time and sample material, and is compatible with high throughput extract analysis, thereby streamlining strain prioritization and evaluation of culturing parameters. With this approach, we analyzed 146 marine Salinispora and Streptomyces strains that were grown and extracted using multiple different protocols. In total, 603 samples were analyzed, generating approximately 1.8 million mass spectra. We constructed a comprehensive molecular network and identified 15 molecular families of diverse natural products and their analogues. The size and breadth of this network shows statistically supported trends in molecular diversity when comparing growth and extraction conditions. The network provides an extensive survey of the biosynthetic capacity of the strain collection and a method to compare strains based on the variety and novelty of their metabolites. This approach allows us to quickly identify patterns in metabolite production that can be linked to taxonomy, culture conditions, and extraction methods, as well as informing the most valuable growth and extraction conditions.

Jordan, PA, Moore BS.  2016.  Biosynthetic pathway connects cryptic ribosomally synthesized posttranslationally modified peptide genes with pyrroloquinoline alkaloids. Cell Chemical Biology. 23:1504-1514.   10.1016/j.chembiol.2016.10.009   AbstractWebsite

In an era where natural product biosynthetic gene clusters can be rapidly identified from sequenced genomes, it is unusual for the biosynthesis of an entire natural product class to remain unknown. Yet, the genetic determinates for pyrroloquinoline alkaloid biosynthesis have remained obscure despite their abundance and deceptive structural simplicity. In this work, we have identified the biosynthetic gene cluster for ammosamides A-C, pyrroloquinoline alkaloids from Streptomyces sp. CNR-698. Through direct cloning, heterologous expression and gene deletions we have validated the ammosamide biosynthetic gene cluster and demonstrated that these seemingly simple molecules are derived from a surprisingly complex set of biosynthetic genes that are also found in the biosynthesis of lymphostin, a structurally related pyrroloquinoline alkaloid from Salinispora and Streptomyces. Our results implicate a conserved set of genes driving pyrroloquinoline biosynthesis that consist of genes frequently associated with ribosomal peptide natural product biosynthesis, and whose exact biochemical role remains enigmatic.

Wang, MX, Carver JJ, Phelan VV, Sanchez LM, Garg N, Peng Y, Nguyen DD, Watrous J, Kapono CA, Luzzatto-Knaan T et al..  2016.  Sharing and community curation of mass spectrometry data with Global Natural Products Social Molecular Networking. Nature Biotechnology. 34:828-837.   10.1038/nbt.3597   AbstractWebsite

The potential of the diverse chemistries present in natural products (NP) for biotechnology and medicine remains untapped because NP databases are not searchable with raw data and the NP community has no way to share data other than in published papers. Although mass spectrometry (MS) techniques are well-suited to high-throughput characterization of NP, there is a pressing need for an infrastructure to enable sharing and curation of data. We present Global Natural Products Social Molecular Networking (GNPS;, an open-access knowledge base for community-wide organization and sharing of raw, processed or identified tandem mass (MS/MS) spectrometry data. In GNPS, crowdsourced curation of freely available community-wide reference MS libraries will underpin improved annotations. Data-driven social-networking should facilitate identification of spectra and foster collaborations. We also introduce the concept of 'living data' through continuous reanalysis of deposited data.

Ross, AC, Gulland LES, Dorrestein PC, Moore BS.  2015.  Targeted capture and heterologous expression of the pseudoalteromonas alterochromide gene cluster in Escherichia coil represents a promising natural product exploratory platform. ACS Synthetic Biology. 4:414-420.   10.1021/sb500280q   AbstractWebsite

Marine pseudoalteromonads represent a very promising source of biologically important natural product molecules. To access and exploit the full chemical capacity of these cosmopolitan Gram-(-) bacteria, we sought to apply universal synthetic biology tools to capture, refactor, and express biosynthetic gene clusters for the production of complex organic compounds in reliable host organisms. Here, we report a platform for the capture of proteobacterial gene dusters using a transformation associated recombination (TAR) strategy coupled with direct pathway manipulation and expression in Escherichia coli. The similar to 34 kb pathway for production of alterochromide lipopeptides by Pseudoalteromonas piscicida JCM 20779 was captured and heterologously expressed in Escherichia coli utilizing native and Escherichia coli-based T7 promoter sequences. Our approach enabled both facile production of the alterochromides and in vivo interrogation of gene function associated with alterochromide's unusual brominated lipid side chain. This platform represents a simple but effective strategy for the discovery and biosynthetic characterization of natural products from marine proteobacteria.

Bonet, B, Teufel R, Crusemann M, Ziemert N, Moore BS.  2015.  Direct capture and heterologous expression of Salinispora natural product genes for the biosynthesis of enterocin. Journal of Natural Products. 78:539-542.   10.1021/np500664q   AbstractWebsite

Heterologous expression of secondary metabolic pathways is a promising approach for the discovery and characterization of bioactive natural products. Herein we report the first heterologous expression of a natural product from the model marine actinomycete genus Salinispora. Using the recently developed method of yeast-mediated transformation-associated recombination for natural product gene clusters, we captured a type II polyketide synthase pathway from Salinispora pacifica with high homology to the enterocin pathway from Streptomyces maritimus and successfully produced enterocin in two different Streptomyces host strains. This result paves the way for the systematic interrogation of Salinispora's promising secondary metabolome.

Mohimani, H, Kersten RD, Liu WT, Wang MX, Purvine SO, Wu S, Brewer HM, Pasa-Tolic L, Bandeira N, Moore BS, Pevzner PA, Dorrestein PC.  2014.  Automated genome mining of ribosomal peptide natural products. Acs Chemical Biology. 9:1545-1551.   10.1021/cb500199h   AbstractWebsite

Ribosomally synthesized and posttranslationally modified peptides (RiPPs), especially from microbial sources, are a large group of bioactive natural products that are a promising source of new (bio)chemistry and bioactivity.(1) In light of exponentially increasing microbial genome databases and improved mass spectrometry (MS)-based metabolornic platforms, there is a need for computational tools that connect natural product genotypes predicted from microbial genome sequences with their corresponding chemotypes from metabolomic data sets. Here, we introduce RiPPquest, a tandem mass spectrometry database search tool for identification of microbial RiPPs, and apply it to lanthipeptide discovery. RiPPquest uses genomics to limit search space to the vicinity of RiPP biosynthetic genes and proteomics to analyze extensive peptide modifications and compute p-values of peptide-spectrum matches (PSMs). We highlight RiPPquest by connecting multiple RiPPs from extracts of Streptomyces to their gene clusters and by the discovery of a new class III lanthipeptide, informatipeptin, from Streptomyces viridochromogenes DSM 40736 to reflect that was discovered by mass spectrometry based genome mining using algorithmic tools rather than manual inspection of mass spectrometry data and genetic information. The presented tool is available at

Nett, M, Moore BS.  2009.  Exploration and engineering of biosynthetic pathways in the marine actinomycete Salinispora tropica. Pure and Applied Chemistry. 81:1075-1084.   10.1351/pac-con-08-08-08   AbstractWebsite

In recent years, members of the marine actinomycete genus Salinispora have proven to be a precious source of structurally diverse secondary metabolites, including the potent anticancer agent salinosporamide A and the enediyne-derived sporolides. The tremendous potential of these marine-dwelling microbes for natural products biosynthesis, however, was not fully realized until sequencing of the Salinispora tropica genome revealed the presence of numerous orphan biosynthetic loci besides a plethora of rare metabolic pathways. This contribution summarizes the biochemical exploration of this prolific organism, highlighting studies in which genome-based information was exploited for the discovery of new enzymatic processes and the engineering of unnatural natural products. Inactivation of key genes within the salinosporamide pathway has expanded its inherent metabolic plasticity and enabled access to various salinosporamide derivatives by mutasynthesis. New insights into the biosynthesis of the sporolides allowed us to increase production titers of these structurally complex molecules, thereby providing the means to search for the DNA cleaving presporolide enediyne.