Genome Mining Tools Research Articles

A chemical ecogenomics approach to understand the roles of secondary metabolites in fungal cereal pathogens.

Secondary metabolites (SMs) are known to play important roles in the virulence and lifestyle of fungal plant pathogens. The increasing availability of fungal pathogen genome sequences and next-generation genomic tools have allowed us to survey the SM gene cluster inventory in individual fungi. Thus, there is immense opportunity for SM discovery in these plant pathogens. Comparative genomics and transcriptomics have been employed to obtain insights on the genetic features that enable fungal pathogens to adapt in individual ecological niches and to adopt the different pathogenic lifestyles. Here, we will discuss how we can use these tools to search for ecologically important SM gene clusters in fungi, using cereal pathogens as models. This ecological genomics approach, combined with genome mining and chemical ecology tools, is likely to advance our understanding of the natural functions of SMs and accelerate bioactive molecule discovery.

Biocatalysts for natural product biosynthesis.

Natural products are important sources of pharmaceuticals, in part owing to their diverse biological activities. Enzymes from natural product biosynthetic pathways have become attractive candidates as biocatalysts for modifying the structures and bioactivities of these complex compounds. Numerous enzymes have been harvested to generate innovative scaffolds, large-scale synthesis of chiral building blocks, and semisynthesis of medicinally relevant natural product derivatives. This review discusses recent examples from three areas: (a) polyketide catalytic domain engineering geared toward synthesis of new polyketides, (b) engineering of tailoring enzymes (other than oxidative enzymes) as biocatalysts, and (c) in vitro total synthesis of natural products using purified enzyme components. With the availability of exponentially increasing genomic information and new genome mining tools, many new and powerful biocatalysts tailored for pharmaceutical synthesis will likely emerge from secondary metabolism.

Natural products genomics

Secondary metabolites (or natural products) are often synthesized by multi‐modular, multi‐domain proteins called non‐ribosomal peptide synthetases (NRPS) and polyketide synthases (PKS). Various well‐known metabolites produced by microorganisms are listed in Table 1, and examples of structures are shown in Fig. 1. In particular, Streptomyces species are known for their ability to produce a wide variety of secondary metabolites such as antibiotics, herbicides, parasitocides, siderophores and pharmacologically active substances including antitumour agents and immunosuppressants. Genome sequencing of Streptomyces coelicolor (Bentley et al., 2002) and S. avermitilis (Omura et al., 2001) revealed over 20 gene clusters for biosynthesis of secondary metabolites, while only a few of their natural products were known prior to sequencing. High‐throughput genome sequencing of hundreds of other bacterial species and strains is now rapidly increasing the repertoire of identified gene clusters for biosynthesis of natural products (Donadio et al., 2007). Here we give a brief update of the current status of genome mining and bioinformatic tools to identify novel NRPS and PKS systems. Table 1 Examples of microbial natural products produced by NRPS/PKS systems. Figure 1 Examples of some chemical structures of (A) polyketides, (B) non‐ribosomal peptides and (C) mixed NRP‐PK compounds. Reprinted with permission from Watanabe and Oikawa (2007). Copyright Royal Society of Chemistry.

Utility of the Trypanosoma cruzi Sequence Database for Identification of Potential Vaccine Candidates by In Silico and In Vitro Screening

Glycosylphosphatidylinositol (GPI)-anchored proteins are abundantly expressed in the infective and intracellular stages of Trypanosoma cruzi and are recognized as antigenic targets by both the humoral and cellular arms of the immune system. Previously, we demonstrated the efficacy of genes encoding GPI-anchored proteins in eliciting partially protective immunity to T. cruzi infection and disease, suggesting their utility as vaccine candidates. For the identification of additional vaccine targets, in this study we screened the T. cruzi expressed sequence tag (EST) and genomic sequence survey (GSS) databases. By applying a variety of web-based genome-mining tools to the analysis of approximately 2,500 sequences, we identified 348 (37.6%) EST and 260 (17.4%) GSS sequences encoding novel parasite-specific proteins. Of these, 19 sequences exhibited the characteristics of secreted and/or membrane-associated GPI proteins. Eight of the selected sequences were amplified to obtain genes TcG1, TcG2, TcG3, TcG4, TcG5, TcG6, TcG7, and TcG8 (TcG1-TcG8) which are expressed in different developmental stages of the parasite and conserved in the genome of a variety of T. cruzi strains. Flow cytometry confirmed the expression of the antigens encoded by the cloned genes as surface proteins in trypomastigote and/or amastigote stages of T. cruzi. When delivered as a DNA vaccine, genes TcG1-TcG6 elicited a parasite-specific antibody response in mice. Except for TcG5, antisera to genes TcG1-TcG6 exhibited trypanolytic activity against the trypomastigote forms of T. cruzi, a property known to correlate with the immune control of T. cruzi. Taken together, our results validate the applicability of bioinformatics in genome mining, resulting in the identification of T. cruzi membrane-associated proteins that are potential vaccine candidates.