Agency: European Commission | Branch: H2020 | Program: RIA | Phase: BIOTEC-03-2016 | Award Amount: 5.06M | Year: 2017
TOPCAPI will exploit the natural fabrication power of actinomycetes as microbial cell factories to produce three high value compounds: GE2270, a starter compound for the semi-synthesis of NAI-Acne, a new topical anti-acne drug in Phase II clinical trials; 6-desmethyl-tetracycline (6DM-TC) and 6-desmethyl 6-deshydro tetracycline (6DM6DH-TC), intermediates for semi-synthetic conversion to medically important type II polyketide tetracyclines (TC), e.g. minocycline, tigecycline, and the novel omadacycline, which is in Phase III clinical trials, to be used against Methicillin-resistant Staphylococcus aureus infections. Our work will focus on two bacterial host species: Streptomyces coelicolor and Streptomyces rimosus. These host species will be characterised using systems biology approaches, applying integrated data analysis to transcriptomics and metabolomics experiments, combined with predictive mathematical modelling to drive the rapid improvement of these microbial cell factories for industrial drug production using advanced metabolic and biosynthetic engineering approaches. At the same time, we will establish an expanded toolbox for the engineering of actinomycete bacteria as cell factories for other high added-value compounds. In the proposed 4-year project, we will: 1. Host engineer two new actinomycete strains for industry-level improved heterologous compound production through integrating systems biology-driven strain design and state-of-the-art genome editing. 2. Engineer the biosynthesis pathways to obtain high-efficiency synthesis of GE2270 and new pathway variants for 6DM-TC and 6DM6DH-TC as well as improve its production purity. 3. Optimise the expression of the engineered target pathways in pre-engineered strains to achieve industrially viable production levels of 1 g/L for GE2270 and 24 g/L for 6DM-TC, while creating a complete novel production strain for 6DM6DH-TC.
Agency: European Commission | Branch: H2020 | Program: RIA | Phase: BIOTEC-03-2016 | Award Amount: 7.48M | Year: 2017
Genetic tractability of bacterial cells allows generating synthetic microbial chassis platforms (SMCPs) with remarkable biotechnological applications but their functionality currently faces important off-genome limitations due to deficient protein-protein interactions, unfavorable protein stoichiometry or generation of toxic intermediates that ultimately compromise the industrial production processes. To solve this problem, Rafts4Biotech project will take advantage of our recent discovery, that bacteria are able to organize subcellular membrane compartments similar to the so-called lipid rafts of eukaryotic cells, to improve/protect specific cellular processes. Rafts4Biotech project will engineer bacterial cells to confine biotechnologically relevant reactions into bacterial lipid rafts to optimize their stoichiometry and protect cells from undesirable metabolic interferences. Hence, the Rafts4Biotech project will produce new generation reliable and robust SMCPs in which industrial production processes are confined in bacterial lipid rafts, released from their classical off-genome limitations and optimized for industrial production. Moreover, this concept can be applied to many prokaryotes, since lipid rafts happens to occur in many bacterial species. Based on this versatility, Raft4Biotech project will use two biotechnologically relevant biosystems, Bacillus subtilis and Escherichia coli, to engineer synthetic bacterial lipid rafts to optimize the performance of three challenging biochemical processes in the fields of pharmaceutical, cosmetics and feed industrial sectors. To achieve this, Rafts4Biotech consortium combines different expertise in synthetic biology, systems biology and mathematical modeling and it includes a number of SMEs that will actively work in this project and will translate this technology into market application. The technology developed by Rafts4Biotec will optimize multistep industrial processes and invigorate European research.
Ortega M.A.,University of Illinois at Urbana - Champaign |
Hao Y.,University of Illinois at Urbana - Champaign |
Walker M.C.,University of Illinois at Urbana - Champaign |
Donadio S.,NAICONS Srl |
And 3 more authors.
Cell Chemical Biology | Year: 2016
Class I lantibiotic dehydratases dehydrate selected Ser/Thr residues of a precursor peptide. Recent studies demonstrated the requirement of glutamyl-tRNAGlu for Ser/Thr activation by one of these enzymes (NisB) from the Firmicute Lactococcus lactis. However, the generality of glutamyl-tRNAGlu usage and the tRNA specificity of lantibiotic dehydratases have not been established. Here we report the 2.7-Å resolution crystal structure, along with the glutamyl-tRNAGlu utilization of MibB, a lantibiotic dehydratase from the Actinobacterium Microbispora sp. 107891 involved in the biosynthesis of the clinical candidate NAI-107. Biochemical assays revealed nucleotides A73 and U72 within the tRNAGlu acceptor stem to be important for MibB glutamyl-tRNAGlu usage. Using this knowledge, an expression system for the production of NAI-107 analogs in Escherichia coli was developed, overcoming the inability of MibB to utilize E. coli tRNAGlu. Our work provides evidence for a common tRNAGlu-dependent dehydration mechanism, paving the way for the characterization of lantibiotics from various phyla. © 2016 Elsevier Ltd. All rights reserved.
Maffioli S.I.,Naicons Srl |
Maffioli S.I.,KtedoGen Srl |
Monciardini P.,Naicons Srl |
Monciardini P.,KtedoGen Srl |
And 9 more authors.
ACS Chemical Biology | Year: 2015
Lantibiotics, an abbreviation for "lanthionine-containing antibiotics", interfere with bacterial metabolism by a mechanism not exploited by the antibiotics currently in clinical use. Thus, they have aroused interest as a source for new therapeutic agents because they can overcome existing resistance mechanisms. Starting from fermentation broth extracts preselected from a high-throughput screening program for discovering cell-wall inhibitors, we isolated a series of related class I lantibiotics produced by different genera of actinomycetes. Analytical techniques together with explorative chemistry have been used to establish their structures: the newly described compounds share a common 24 aa sequence with the previously reported lantibiotic planosporicin (aka 97518), differing at positions 4, 6, and 14. All of these compounds maintain an overall -1 charge at physiological pH. While all of these lantibiotics display modest antibacterial activity, their potency can be substantially modulated by progressively eliminating the negative charges, with the most active compounds carrying basic amide derivatives of the two carboxylates originally present in the natural compounds. Interestingly, both natural and chemically modified lantibiotics target the key biosynthetic intermediate lipid II, but the former compounds do not bind as effectively as the latter in vivo. Remarkably, the basic derivatives display an antibacterial potency and a killing effect similar to those of NAI-107, a distantly related actinomycete-produced class I lantibiotic which lacks altogether carboxyl groups and which is a promising clinical candidate for treating Gram-positive infections caused by multi-drug-resistant pathogens. © 2015 American Chemical Society.
Zhang Y.,Rutgers University |
Degen D.,Rutgers University |
Ho M.X.,Rutgers University |
Sineva E.,Rutgers University |
And 13 more authors.
eLife | Year: 2014
Using a combination of genetic, biochemical, and structural approaches, we show that the cyclic-peptide antibiotic GE23077 (GE) binds directly to the bacterial RNA polymerase (RNAP) active-center 'i' and 'i+1' nucleotide binding sites, preventing the binding of initiating nucleotides, and thereby preventing transcription initiation. The target-based resistance spectrum for GE is unusually small, reflecting the fact that the GE binding site on RNAP includes residues of the RNAP active center that cannot be substituted without loss of RNAP activity. The GE binding site on RNAP is different from the rifamycin binding site. Accordingly, GE and rifamycins do not exhibit cross-resistance, and GE and a rifamycin can bind simultaneously to RNAP. The GE binding site on RNAP is immediately adjacent to the rifamycin binding site. Accordingly, covalent linkage of GE to a rifamycin provides a bipartite inhibitor having very high potency and very low susceptibility to target-based resistance. © Zhang et al.
Agency: European Commission | Branch: H2020 | Program: MSCA-ITN-ETN | Phase: MSCA-ITN-2016 | Award Amount: 3.93M | Year: 2017
Train2Target is a multidisciplinary European Training Network built to address the challenge of the discovery of alternative antimicrobials. Innovative strategies to deliver a next generation of drugs are urgently needed. The alarming threats and spread of multi-drug resistant bacteria is currently leaving clinicians with very limited options to combat infections especially those from Gram-negative pathogens. The Train 2Target research programme focuses on the assembly of the well-known bacterial cell envelope from a new perspective. Indeed it aims to inhibit novel targets in envelope biogenesis by altering the function and misbalancing the coordination of envelope assembly machines, which build and assemble the Gram-negative bacterial envelope. A wide variety of chemical classes and compounds sources will be screened using innovative biochemical, biophysical and genetic assays to identify valuable hit scaffolds to be optimized into druggable leads. The high quality and credibility of our consortium is ensured by a strong interdisciplinary academia-industry partnership to encompass different complementary expertise ranging from microbiology, bacterial genetics, biochemistry, cell imaging, structural biology, biophysics and chemical synthesis. Our 9 academic groups are all renowned leaders in the cell envelope biogenesis field, whereas the complementary 5 SMEs and 3 Industry partners are specialised in drug discovery and development of novel anti-infective drugs. This unique combination of scientific excellence and industrial know-how in drug discovery covers the entire process from the design to the implementation of innovative antibacterial strategies and lead identification. Train2Target also represents a unique research platform to train 15 Early Stage Researchers and equip them with the necessary scientific and transferable skills that will make them highly competitive for both top European research institutions and the pharma/biotech job market.
Iorio M.,NAICONS Srl |
Sasso O.,Italian Institute of Technology |
Maffioli S.I.,NAICONS Srl |
Bertorelli R.,Italian Institute of Technology |
And 11 more authors.
ACS Chemical Biology | Year: 2014
Among the growing family of ribosomally synthesized, post-translationally modified peptides, particularly intriguing are class III lanthipeptides containing the triamino acid labionin. In the course of a screening program aimed at finding bacterial cell wall inhibitors, we discovered a new lanthipeptide produced by an Actinoplanes sp. The molecule, designated NAI-112, consists of 22 amino acids and contains an N-terminal labionin and a C-terminal methyl-labionin. Unique among lanthipeptides, it carries a 6-deoxyhexose moiety N-linked to a tryptophan residue. Consistently, the corresponding gene cluster encodes, in addition to the LanKC enzyme characteristic of this lanthipeptide class, a glycosyl transferase. Despite possessing weak antibacterial activity, NAI-112 is effective in experimental models of nociceptive pain, reducing pain symptoms in mice in both the formalin and the chronic constriction injury tests. Thus, NAI-112 represents, after the labyrinthopeptins, the second example of a lanthipeptide effective against nociceptive pain. © 2013 American Chemical Society.
Lo Grasso L.,University of Palermo |
Maffioli S.,Naicons Srl |
Sosio M.,Naicons Srl |
Bib M.,John Innes Center |
And 2 more authors.
Journal of Bacteriology | Year: 2015
The actinomycete Nonomuraea sp. strain ATCC 39727 produces the glycopeptide A40926, the precursor of dalbavancin. Biosynthesis of A40926 is encoded by the dbv gene cluster, which contains 37 protein-coding sequences that participate in antibiotic biosynthesis, regulation, immunity, and export. In addition to the positive regulatory protein Dbv4, the A40926-biosynthetic gene cluster encodes two additional putative regulators, Dbv3 and Dbv6. Independent mutations in these genes, combined with bioassays and liquid chromatography-mass spectrometry (LC-MS) analyses, demonstrated that Dbv3 and Dbv4 are both required for antibiotic production, while inactivation of dbv6 had no effect. In addition, overexpression of dbv3 led to higher levels of A40926 production. Transcriptional and quantitative reverse transcription (RT)-PCR analyses showed that Dbv4 is essential for the transcription of two operons, dbv14-dbv8 and dbv30-dbv35, while Dbv3 positively controls the expression of four monocistronic transcription units (dbv4, dbv29, dbv36, and dbv37) and of six operons (dbv2-dbv1, dbv14-dbv8, dbv17-dbv15, dbv21- dbv20, dbv24-dbv28, and dbv30-dbv35). We propose a complex and coordinated model of regulation in which Dbv3 directly or indirectly activates transcription of dbv4 and controls biosynthesis of 4-hydroxyphenylglycine and the heptapeptide backbone, A40926 export, and some tailoring reactions (mannosylation and hexose oxidation), while Dbv4 directly regulates biosynthesis of 3,5-dihydroxyphenylglycine and other tailoring reactions, including the four cross-links, halogenation, glycosylation, and acylation. © 2015, American Society for Microbiology.
Monciardini P.,Naicons Srl |
Iorio M.,Naicons Srl |
Maffioli S.,Naicons Srl |
Sosio M.,Naicons Srl |
Donadio S.,Naicons Srl
Microbial Biotechnology | Year: 2014
There is an increased need for new drug leads to treat diseases in humans, animals and plants. A dramatic example is represented by the need for novel and more effective antibiotics to combat multidrug-resistant microbial pathogens. Natural products represent a major source of approved drugs and still play an important role in supplying chemical diversity, despite a decreased interest by large pharmaceutical companies. Novel approaches must be implemented to decrease the chances of rediscovering the tens of thousands of known natural products. In this review, we present an overview of natural product screening, focusing particularly on microbial products. Different approaches can be implemented to increase the probability of finding new bioactive molecules. We thus present the rationale and selected examples of the use of hypersensitive assays; of accessing unexplored microorganisms, including the metagenome; and of genome mining. We then focus our attention on the technology platform that we are currently using, consisting of approximately 70000 microbial strains, mostly actinomycetes and filamentous fungi, and discuss about high-quality screening in the search for bioactive molecules. Finally, two case studies are discussed, including the spark that arose interest in the compound: in the case of orthoformimycin, the novel mechanism of action predicted a novel structural class; in the case of NAI-112, structural similarity pointed out to a possible in vivo activity. Both predictions were then experimentally confirmed. © 2014 The Authors. Microbial Biotechnology published by John Wiley & Sons Ltd and Society for Applied Microbiology.
Naicons Srl | Date: 2013-11-27
The present invention concerns novel antibiotic compounds, which are lantibiotics, the processes for their preparation, their pharmaceutically acceptable salts, pharmaceutical compositions containing the lantibiotics, and their use as antibacterial agents. Compounds designated as lantibiotics, such as those of the present invention, are peptides belonging to the general class of antibiotic compounds, and are further generally characterized by the presence of the amino acids lanthionine and/or 3-methyllanthionine. The novel lantibiotic compounds are active against bacterial infections caused by Clostridium difficile, Staphylococcus spp., Streptococcus spp, Enterococcus spp., and other bacteria.