Eric W. Schmidt

Ribosomally synthesized and post-translationally modified peptide natural products: overview and recommendations for a universal nomenclature

This review presents recommended nomenclature for the biosynthesis of ribosomally synthesized and post-translationally modified peptides (RiPPs), a rapidly growing class of natural products. The current knowledge regarding the biosynthesis of the >20 distinct compound classes is also reviewed, and commonalities are discussed.

A global assembly line for cyanobactins.

More than 100 cyclic peptides harboring heterocyclized residues are known from marine ascidians, sponges and different genera of cyanobacteria. Here, we report an assembly line responsible for the biosynthesis of these diverse peptides, now called cyanobactins, both in symbiotic and free-living cyanobacteria. By comparing five new cyanobactin biosynthetic clusters, we produced the prenylated antitumor preclinical candidate trunkamide in Escherichia coli culture using genetic engineering.

Ribosomal peptide natural products: bridging the ribosomal and nonribosomal worlds.

Ribosomally synthesized bacterial natural products rival the nonribosomal peptides in their structural and functional diversity. The last decade has seen substantial progress in the identification and characterization of biosynthetic pathways leading to ribosomal peptide natural products with new and unusual structural motifs. In some of these cases, the motifs are similar to those found in nonribosomal peptides, and many are constructed by convergent or even paralogous enzymes. Here, we summarize the major structural and biosynthetic categories of ribosomally synthesized bacterial natural products and, where applicable, compare them to their homologs from nonribosomal biosynthesis.

Structure of Trichamide, a Cyclic Peptide from the Bloom-Forming Cyanobacterium Trichodesmium erythraeum, Predicted from the Genome Sequence

ABSTRACT A gene cluster for the biosynthesis of a new small cyclic peptide, dubbed trichamide, was discovered in the genome of the global, bloom-forming marine cyanobacterium Trichodesmium erythraeum ISM101 because of striking similarities to the previously characterized patellamide biosynthesis cluster. The tri cluster consists of a precursor peptide gene containing the amino acid sequence for mature trichamide, a putative heterocyclization gene, an oxidase, two proteases, and hypothetical genes. Based upon detailed sequence analysis, a structure was predicted for trichamide and confirmed by Fourier transform mass spectrometry. Trichamide consists of 11 amino acids, including two cysteine-derived thiazole groups, and is cyclized by an N—C terminal amide bond. As the first natural product reported from T. erythraeum, trichamide shows the power of genome mining in the prediction and discovery of new natural products.

Metagenomic Approaches to Natural Products from Free-Living and Symbiotic Organisms

Bacterial cultivation has been a mainstay of natural products discovery for the past 80 years. However, the majority of bacteria are recalcitrant to culture, providing an untapped source for new natural products. Metagenomic analysis provides an alternative method to directly access the uncultivated genome for natural products research and for the discovery of novel, bioactive substances. Applications of metagenomics to diverse habitats, such as soils and the interior of animals, are described.

Complex microbiome underlying secondary and primary metabolism in the tunicate-Prochloron symbiosis

The relationship between tunicates and the uncultivated cyanobacterium Prochloron didemni has long provided a model symbiosis. P. didemni is required for survival of animals such as Lissoclinum patella and also makes secondary metabolites of pharmaceutical interest. Here, we present the metagenomes, chemistry, and microbiomes of four related L. patella tunicate samples from a wide geographical range of the tropical Pacific. The remarkably similar P. didemni genomes are the most complex so far assembled from uncultivated organisms. Although P. didemni has not been stably cultivated and comprises a single strain in each sample, a complete set of metabolic genes indicates that the bacteria are likely capable of reproducing outside the host. The sequences reveal notable peculiarities of the photosynthetic apparatus and explain the basis of nutrient exchange underlying the symbiosis. P. didemni likely profoundly influences the lipid composition of the animals by synthesizing sterols and an unusual lipid with biofuel potential. In addition, L. patella also harbors a great variety of other bacterial groups that contribute nutritional and secondary metabolic products to the symbiosis. These bacteria possess an enormous genetic potential to synthesize new secondary metabolites. For example, an antitumor candidate molecule, patellazole, is not encoded in the genome of Prochloron and was linked to other bacteria from the microbiome. This study unveils the complex L. patella microbiome and its impact on primary and secondary metabolism, revealing a remarkable versatility in creating and exchanging small molecules.

Using Marine Natural Products to Discover a Protease that Catalyzes Peptide Macrocyclization of Diverse Substrates

Small N-C terminally cyclic ribosomal peptides are common in nature, yet the mechanisms underlying their biosyntheses remain largely unknown. We recently identified candidate peptide cyclase genes in the metagenomes of marine animals. Here we report that PatG, a protease discovered in this analysis, cleaves variables, short peptides out of a precursor protein, and cyclizes these peptides in vivo and in vitro.

Thioesterase-Like Role for Fungal PKS-NRPS Hybrid Reductive Domains

Fungal reduced polyketides possess diverse structures exploring a broad region of chemical space despite their synthesis by very similar enzymes. Many fungal polyketides are capped by diverse amino acid-derived five-membered rings, the tetramic acids and related pyrrolidine-2-ones. The known tetramic acid synthetase enzymes in fungi contain C-terminal reductive (R) domains that were proposed to release reduced pyrrolidine-2-one intermediates en route to the tetramic acids. To determine the enzymatic basis of pyrrolidine-2-one diversity, we overexpressed equisetin synthetase (EqiS) R domains and analyzed their reactivity with synthetic substrate analogs. We show that the EqiS R domain does not perform a reducing function and does not bind reducing cofactors. Instead, the EqiS R catalyzes a Dieckmann condensation, with an estimated kcat approximately 15 s(-1). This role differs from the redox reactions normally catalyzed by short chain dehydrogenase/reductase superfamily enzymes.

Genome streamlining and chemical defense in a coral reef symbiosis

Secondary metabolites are ubiquitous in bacteria, but by definition, they are thought to be nonessential. Highly toxic secondary metabolites such as patellazoles have been isolated from marine tunicates, where their exceptional potency and abundance implies a role in chemical defense, but their biological source is unknown. Here, we describe the association of the tunicate Lissoclinum patella with a symbiotic α-proteobacterium, Candidatus Endolissoclinum faulkneri, and present chemical and biological evidence that the bacterium synthesizes patellazoles. We sequenced and assembled the complete Ca. E. faulkneri genome, directly from metagenomic DNA obtained from the tunicate, where it accounted for 0.6% of sequence data. We show that the large patellazoles biosynthetic pathway is maintained, whereas the remainder of the genome is undergoing extensive streamlining to eliminate unneeded genes. The preservation of this pathway in streamlined bacteria demonstrates that secondary metabolism is an essential component of the symbiotic interaction.

The complete genome of Teredinibacter turnerae T7901: An intracellular endosymbiont of marine wood-boring bivalves (shipworms)

Here we report the complete genome sequence of Teredinibacter turnerae T7901. T. turnerae is a marine gamma proteobacterium that occurs as an intracellular endosymbiont in the gills of wood-boring marine bivalves of the family Teredinidae (shipworms). This species is the sole cultivated member of an endosymbiotic consortium thought to provide the host with enzymes, including cellulases and nitrogenase, critical for digestion of wood and supplementation of the hosts nitrogen-deficient diet. T. turnerae is closely related to the free-living marine polysaccharide degrading bacterium Saccharophagus degradans str. 2–40 and to as yet uncultivated endosymbionts with which it coexists in shipworm cells. Like S. degradans, the T. turnerae genome encodes a large number of enzymes predicted to be involved in complex polysaccharide degradation (>100). However, unlike S. degradans, which degrades a broad spectrum (>10 classes) of complex plant, fungal and algal polysaccharides, T. turnerae primarily encodes enzymes associated with deconstruction of terrestrial woody plant material. Also unlike S. degradans and many other eubacteria, T. turnerae dedicates a large proportion of its genome to genes predicted to function in secondary metabolism. Despite its intracellular niche, the T. turnerae genome lacks many features associated with obligate intracellular existence (e.g. reduced genome size, reduced %G+C, loss of genes of core metabolism) and displays evidence of adaptations common to free-living bacteria (e.g. defense against bacteriophage infection). These results suggest that T. turnerae is likely a facultative intracellular ensosymbiont whose niche presently includes, or recently included, free-living existence. As such, the T. turnerae genome provides insights into the range of genomic adaptations associated with intracellular endosymbiosis as well as enzymatic mechanisms relevant to the recycling of plant materials in marine environments and the production of cellulose-derived biofuels.