Harnessing fungal nonribosomal cyclodepsipeptide synthetases for mechanistic insights and tailored engineering

Steiniger, Charlotte; Hoffmann, Sylvester; Mainz, Andi; Kaiser, Marcel; Voigt, Kerstin; Meyer, Vera; Süssmuth, Roderich D.

doi:10.1039/c7sc03093b

Cited by 38 publications

(88 citation statements)

References 37 publications

(74 reference statements)

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“…Using A-domains which accept methylated amino acids in domain swaps could also be used to introduce methylated amino acids, or unnatural analogues such as the fluorinated amino acids into other NRP natural products of agricultural of pharmaceutical interest. Indeed, there have been some major advances in NRPS structural biology and engineering recently, 21 – 23 which means that the prospect of swapping in domains/modules to introduce N -methylated amino acids into existing NRPs could become feasible. Detailed in vitro studies of the N -MeT enzyme and the N –Me amino acid activating A-domains and modules, including crystallography, could help to elucidate where the selectivity and specificity of such systems lie, and the roles that the individual NRPS domains play in controlling the output of such systems.…”

Section: Discussionmentioning

confidence: 99%

The cycloaspeptides: uncovering a new model for methylated nonribosomal peptide biosynthesis

et al. 2018

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Section: Discussionmentioning

confidence: 99%

The cycloaspeptides: uncovering a new model for methylated nonribosomal peptide biosynthesis

et al. 2018

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“…Likewise, cyclic orbitide peptides from Euphorbiaceae spp. and novel cyclodepsipeptides of fungal organisms represent intriguing antifungal templates [ 187 , 188 ]. Indeed, naturally occurring peptide-based molecules have illustrated success in this regard and are approved for clinical use as antifungals.…”

Section: Development Of Peptide Anti-infectives Targeting Fungal Rmentioning

confidence: 99%

Regulated Cell Death as a Therapeutic Target for Novel Antifungal Peptides and Biologics

Yeaman

Büttner

Thevissen

2018

Oxidative Medicine and Cellular Longevity

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The rise of microbial pathogens refractory to conventional antibiotics represents one of the most urgent and global public health concerns for the 21st century. Emergence of Candida auris isolates and the persistence of invasive mold infections that resist existing treatment and cause severe illness has underscored the threat of drug-resistant fungal infections. To meet these growing challenges, mechanistically novel agents and strategies are needed that surpass the conventional fungistatic or fungicidal drug actions. Host defense peptides have long been misunderstood as indiscriminant membrane detergents. However, evidence gathered over the past decade clearly points to their sophisticated and selective mechanisms of action, including exploiting regulated cell death pathways of their target pathogens. Such peptides perturb transmembrane potential and mitochondrial energetics, inducing phosphatidylserine accessibility and metacaspase activation in fungi. These mechanisms are often multimodal, affording target pathogens fewer resistance options as compared to traditional small molecule drugs. Here, recent advances in the field are examined regarding regulated cell death subroutines as potential therapeutic targets for innovative anti-infective peptides against pathogenic fungi. Furthering knowledge of protective host defense peptide interactions with target pathogens is key to advancing and applying novel prophylactic and therapeutic countermeasures to fungal resistance and pathogenesis.

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“…Ester bond formation as well as macrocyclization occurs at the C 3 domain. However, based on recent results by Yu et al [ 11 ] and from our group [ 12 ], experimental evidence points to the “linear” or “looping” model: the elongation occurs by the attachment of a single building block (hydroxy acid or N -methyl amino acid), while the growing depsipeptide chain is passed between the PCP 1 and the PCP 2a/b domains. Peptide bond formation is catalysed in the C 2 domain, while the C 3 domain catalyses ester bond formation and macrocyclization.…”

Section: Introductionmentioning

confidence: 99%

“…Peptide bond formation is catalysed in the C 2 domain, while the C 3 domain catalyses ester bond formation and macrocyclization. In this model, the role of the double PCP 2a/b domains remains unclear, as either one of the domains is sufficient for biosynthesis of the final product [ 11 , 12 ]. It was proposed that C 1 has no direct catalytic function, because truncated CDP synthetases missing the C 1 domain are still functional.…”

Section: Introductionmentioning

confidence: 99%

“…It was proposed that C 1 has no direct catalytic function, because truncated CDP synthetases missing the C 1 domain are still functional. Thus, C 1 could rather have a stabilizing or supportive role during the catalytic cycle [ 12 ]. Currently, it is not clear which of these two models accurately represents NRPs activity, and more investigations are required to fully understand the underlying mechanism of CDP biosynthesis.…”

Section: Introductionmentioning

confidence: 99%

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Aspergillus niger is a superior expression host for the production of bioactive fungal cyclodepsipeptides

Boecker

Gratz

Kerwat

et al. 2018

Fungal Biol Biotechnol

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BackgroundFungal cyclodepsipeptides (CDPs) are non-ribosomally synthesized peptides produced by a variety of filamentous fungi and are of interest to the pharmaceutical industry due to their anticancer, antimicrobial and anthelmintic bioactivities. However, both chemical synthesis and isolation of CDPs from their natural producers are limited due to high costs and comparatively low yields. These challenges might be overcome by heterologous expression of the respective CDP-synthesizing genes in a suitable fungal host. The well-established industrial fungus Aspergillus niger was recently genetically reprogrammed to overproduce the cyclodepsipeptide enniatin B in g/L scale, suggesting that it can generally serve as a high production strain for natural products such as CDPs. In this study, we thus aimed to determine whether other CDPs such as beauvericin and bassianolide can be produced with high titres in A. niger, and whether the generated expression strains can be used to synthesize new-to-nature CDP derivatives.ResultsThe beauvericin and bassianolide synthetases were expressed under control of the tuneable Tet-on promoter, and titres of about 350–600 mg/L for bassianolide and beauvericin were achieved when using optimized feeding conditions, respectively. These are the highest concentrations ever reported for both compounds, whether isolated from natural or heterologous expression systems. We also show that the newly established Tet-on based expression strains can be used to produce new-to-nature beauvericin derivatives by precursor directed biosynthesis, including the compounds 12-hydroxyvalerate-beauvericin and bromo-beauvericin. By feeding deuterated variants of one of the necessary precursors (d-hydroxyisovalerate), we were able to purify deuterated analogues of beauvericin and bassianolide from the respective A. niger expression strains. These deuterated compounds could potentially be used as internal standards in stable isotope dilution analyses to evaluate and quantify fungal spoilage of food and feed products.ConclusionIn this study, we show that the product portfolio of A. niger can be expanded from enniatin to other CDPs such as beauvericin and bassianolide, as well as derivatives thereof. This illustrates the capability of A. niger to produce a range of different peptide natural products in titres high enough to become industrially relevant.Electronic supplementary materialThe online version of this article (10.1186/s40694-018-0048-3) contains supplementary material, which is available to authorized users.

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Harnessing fungal nonribosomal cyclodepsipeptide synthetases for mechanistic insights and tailored engineering

Abstract: Hybrid fungal CDP synthetases are constructed from three different origins to produce highly active cyclodepsipeptides up to g L–1 scale.

Cited by 38 publications

References 37 publications

The cycloaspeptides: uncovering a new model for methylated nonribosomal peptide biosynthesis

The cycloaspeptides: uncovering a new model for methylated nonribosomal peptide biosynthesis

Regulated Cell Death as a Therapeutic Target for Novel Antifungal Peptides and Biologics

Aspergillus niger is a superior expression host for the production of bioactive fungal cyclodepsipeptides

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