Antibiotic drug discovery Wohlleben, Wolfgang; Mast, Yvonne; Stegmann, Evi ...
Microbial biotechnology,
September 2016, Letnik:
9, Številka:
5
Journal Article
Recenzirano
Odprti dostop
Summary
Due to the threat posed by the increase of highly resistant pathogenic bacteria, there is an urgent need for new antibiotics; all the more so since in the last 20 years, the approval for new ...antibacterial agents had decreased. The field of natural product discovery has undergone a tremendous development over the past few years. This has been the consequence of several new and revolutionizing drug discovery and development techniques, which is initiating a ‘New Age of Antibiotic Discovery’. In this review, we concentrate on the most significant discovery approaches during the last and present years and comment on the challenges facing the community in the coming years.
The field of natural product discovery has undergone a tremendous development over the past few years. This has been the consequence of several new and revolutionizing drug discovery and development techniques. In this review, we concentrate on the most significant discovery approaches in the last and present years and comment on the challenges facing the community in the coming years.
Nitrogen is an essential element required for bacterial growth. It serves as a building block for the biosynthesis of macromolecules and provides precursors for secondary metabolites. Bacteria have ...developed the ability to use various nitrogen sources and possess two enzyme systems for nitrogen assimilation involving glutamine synthetase/glutamate synthase and glutamate dehydrogenase. Microorganisms living in habitats with changeable availability of nutrients have developed strategies to survive under nitrogen limitation. One adaptation is the ability to acquire nitrogen from alternative sources including the polyamines putrescine, cadaverine, spermidine and spermine, as well as the monoamine ethanolamine. Bacterial polyamine and monoamine metabolism is not only important under low nitrogen availability, but it is also required to survive under high concentrations of these compounds. Such conditions can occur in diverse habitats such as soil, plant tissues and human cells. Strategies of pathogenic and non-pathogenic bacteria to survive in the presence of poly- and monoamines offer the possibility to combat pathogens by using their capability to metabolize polyamines as an antibiotic drug target. This work aims to summarize the knowledge on poly- and monoamine metabolism in bacteria and its role in nitrogen metabolism.
Lanthipeptides are a class of ribosomally synthesised and post-translationally modified peptide (RiPP) natural products from the bacterial secondary metabolism. Their name is derived from the ...characteristic lanthionine or methyl-lanthionine residues contained in the processed peptide. Lanthipeptides that possess an antibacterial activity are called lantibiotics. Whereas multiple tools exist to identify lanthipeptide gene clusters from genomic data, no programs are available to predict the post-translational modifications of lanthipeptides, such as the proteolytic cleavage of the leader peptide part or tailoring modifications based on the analysis of the gene cluster sequence. antiSMASH is a software pipeline for the identification of secondary metabolite biosynthetic clusters from genomic input and the prediction of products produced by the identified clusters. Here we present a novel antiSMASH module using a rule-based approach to combine signature motifs for biosynthetic enzymes and lanthipeptide-specific cleavage site motifs to identify lanthipeptide clusters in genomic data, assign the specific lanthipeptide class, predict prepeptide cleavage, tailoring reactions, and the processed molecular weight of the mature peptide products.
Abstract
Streptomycetes have to cope with fluctuating nitrogen conditions in soil. Key enzymes in nitrogen metabolism in streptomycetes are the two glutamine synthetases (GSs), GlnA and GlnII. We ...demonstrated that GlnA3 and GlnA4, proteins previously annotated as GSs, are involved in utilization and detoxification of polyamines and ethanolamine. Homologs of these enzymes are present in pathogenic Actinobacteria and are promising drug targets. We designed a novel strategy for development of anti-tubercular antibiotics based on the inhibition of GlnA3.
Streptomyces coelicolor GlnR is a global regulator that controls genes involved in nitrogen metabolism. By genomic screening 10 new GlnR targets were identified, including enzymes for ammonium ...assimilation (glnII, gdhA), nitrite reduction (nirB), urea cleavage (ureA) and a number of biochemically uncharacterized proteins (SCO0255, SCO0888, SCO2195, SCO2400, SCO2404, SCO7155). For the GlnR regulon, a GlnR binding site which comprises the sequence gTnAc-n₆-GaAAc-n₆-GtnAC-n₆-GAAAc-n₆ has been found. Reverse transcription analysis of S. coelicolor and the S. coelicolor glnR mutant revealed that GlnR activates or represses the expression of its target genes. Furthermore, glnR expression itself was shown to be nitrogen-dependent. Physiological studies of S. coelicolor and the S. coelicolor glnR mutant with ammonium and nitrate as the sole nitrogen source revealed that GlnR is not only involved in ammonium assimilation but also in ammonium supply. blast analysis demonstrated that GlnR-homologous proteins are present in different actinomycetes containing the glnA gene with the conserved GlnR binding site. By DNA binding studies, it was furthermore demonstrated that S. coelicolor GlnR is able to interact with these glnA upstream regions. We therefore suggest that GlnR-mediated regulation is not restricted to Streptomyces but constitutes a regulon conserved in many actinomycetes.
Non-ribosomal peptide synthetases (NRPSs) are large multimodular enzymes that synthesize a wide range of biologically active natural peptide compounds, of which many are pharmacologically important. ...Peptide bond formation is catalyzed by the Condensation (C) domain. Various functional subtypes of the C domain exist: An LCL domain catalyzes a peptide bond between two L-amino acids, a DCL domain links an L-amino acid to a growing peptide ending with a D-amino acid, a Starter C domain (first denominated and classified as a separate subtype here) acylates the first amino acid with a beta-hydroxy-carboxylic acid (typically a beta-hydroxyl fatty acid), and Heterocyclization (Cyc) domains catalyze both peptide bond formation and subsequent cyclization of cysteine, serine or threonine residues. The homologous Epimerization (E) domain flips the chirality of the last amino acid in the growing peptide; Dual E/C domains catalyze both epimerization and condensation.
In this paper, we report on the reconstruction of the phylogenetic relationship of NRPS C domain subtypes and analyze in detail the sequence motifs of recently discovered subtypes (Dual E/C, DCL and Starter domains) and their characteristic sequence differences, mutually and in comparison with LCL domains. Based on their phylogeny and the comparison of their sequence motifs, LCL and Starter domains appear to be more closely related to each other than to other subtypes, though pronounced differences in some segments of the protein account for the unequal donor substrates (amino vs. beta-hydroxy-carboxylic acid). Furthermore, on the basis of phylogeny and the comparison of sequence motifs, we conclude that Dual E/C and DCL domains share a common ancestor. In the same way, the evolutionary origin of a C domain of unknown function in glycopeptide (GP) NRPSs can be determined to be an LCL domain. In the case of two GP C domains which are most similar to DCL but which have LCL activity, we postulate convergent evolution.
We systematize all C domain subtypes including the novel Starter C domain. With our results, it will be easier to decide the subtype of unknown C domains as we provide profile Hidden Markov Models (pHMMs) for the sequence motifs as well as for the entire sequences. The determined specificity conferring positions will be helpful for the mutation of one subtype into another, e.g. turning DCL to LCL, which can be a useful step for obtaining novel products.
Polyketides belong to the most valuable natural products, including diverse bioactive compounds, such as antibiotics, anticancer drugs, antifungal agents, immunosuppressants and others. Their ...structures are assembled by polyketide synthases (PKSs). Modular PKSs are composed of modules, which involve sets of domains catalysing the stepwise polyketide biosynthesis. The acyltransferase (AT) domains and their "partners", the acyl carrier proteins (ACPs), thereby play an essential role. The AT loads the building blocks onto the "substrate acceptor", the ACP. Thus, the AT dictates which building blocks are incorporated into the polyketide structure. The precursor- and occasionally the ACP-specificity of the ATs differ across the polyketide pathways and therefore, the ATs contribute to the structural diversity within this group of complex natural products. Those features make the AT enzymes one of the most promising tools for manipulation of polyketide assembly lines and generation of new polyketide compounds. However, the AT-based PKS engineering is still not straightforward and thus, rational design of functional PKSs requires detailed understanding of the complex machineries. This review summarizes the attempts of PKS engineering by exploiting the AT attributes for the modification of polyketide structures. The article includes 253 references and covers the most relevant literature published until May 2018.
By culturing microorganisms under standard laboratory conditions, most biosynthetic gene clusters (BGCs) are not expressed, and thus, the products are not produced. To explore this biosynthetic ...potential, we developed a novel “semi-targeted” approach focusing on activating “silent” BGCs by concurrently introducing a group of regulator genes into streptomycetes of the Tübingen strain collection. We constructed integrative plasmids containing two classes of regulatory genes under the control of the constitutive promoter ermE*p (cluster situated regulators (CSR) and Streptomyces antibiotic regulatory proteins (SARPs)). These plasmids were introduced into Streptomyces sp. TÜ17, Streptomyces sp. TÜ10 and Streptomyces sp. TÜ102. Introduction of the CSRs-plasmid into strain S. sp. TÜ17 activated the production of mayamycin A. By using the individual regulator genes, we proved that Aur1P, was responsible for the activation. In strain S. sp. TÜ102, the introduction of the SARP-plasmid triggered the production of a chartreusin-like compound. Insertion of the CSRs-plasmid into strain S. sp. TÜ10 resulted in activating the warkmycin-BGC. In both recombinants, activation of the BGCs was only possible through the simultaneous expression of aur1PR3 and griR in S. sp. TÜ102 and aur1P and pntR in of S. sp. TÜ10.
is a soil bacterium living in a habitat with very changeable nutrient availability. This organism possesses a complex nitrogen metabolism and is able to utilize the polyamines putrescine, cadaverine, ...spermidine, and spermine and the monoamine ethanolamine. We demonstrated that GlnA2 (SCO2241) facilitates
to survive under high toxic polyamine concentrations. GlnA2 is a gamma-glutamylpolyamine synthetase, an enzyme catalyzing the first step in polyamine catabolism. The role of GlnA2 was confirmed in phenotypical studies with a
deletion mutant as well as in transcriptional and biochemical analyses. Among all GS-like enzymes in
, GlnA2 possesses the highest specificity towards short-chain polyamines (putrescine and cadaverine), while its functional homolog GlnA3 (SCO6962) prefers long-chain polyamines (spermidine and spermine) and GlnA4 (SCO1613) accepts only monoamines. The genome-wide RNAseq analysis in the presence of the polyamines putrescine, cadaverine, spermidine, or spermine revealed indication of the occurrence of different routes for polyamine catabolism in
involving GlnA2 and GlnA3. Furthermore, GlnA2 and GlnA3 are differently regulated. From our results, we can propose a complemented model of polyamine catabolism in
, which involves the gamma-glutamylation pathway as well as other alternative utilization pathways.
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