Towards a sustainable generation of pseudopterosin-type bioactives

Ringel, Marion; Reinbold, Markus; Hirte, Max; Haack, Martina; Huber, Claudia; Eisenreich, Wolfgang; Masri, Mahmoud; Schenk, Gerhard; Guddat, Luke W.; Loll, Bernhard; Kerr, Russell G.; Garbe, Daniel; Brück, Thomas

doi:10.1039/d0gc01697g

Cited by 10 publications

(9 citation statements)

References 78 publications

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“…A pH-based feeding protocol was set as previously described [ 19 , 21 ]. The feeding solution consisted of 600 g l −1 glycerol, 5 g l −1 yeast extract, 35 g l −1 collagen, 20 g l −1 MgSO4, 0.3 g l −1 thiamine-HCl, 5 ml l −1 1 M ammonium iron(III) citrate, 20 ml l −1 100 × trace element solution (pH = 7.0) [ 19 , 47 ].…”

Section: Methodsmentioning

confidence: 99%

Biotechnological potential and initial characterization of two novel sesquiterpene synthases from Basidiomycota Coniophora puteana for heterologous production of δ-cadinol

et al. 2022

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Background Terpene synthases are versatile catalysts in all domains of life, catalyzing the formation of an enormous variety of different terpenoid secondary metabolites. Due to their diverse bioactive properties, terpenoids are of great interest as innovative ingredients in pharmaceutical and cosmetic applications. Recent advances in genome sequencing have led to the discovery of numerous terpene synthases, in particular in Basidiomycota like the wood rotting fungus Coniophora puteana, which further enhances the scope for the manufacture of terpenes for industrial purposes. Results In this study we describe the identification of two novel (+)-δ-cadinol synthases from C. puteana, Copu5 and Copu9. The sesquiterpene (+)-δ-cadinol was previously shown to exhibit cytotoxic activity therefore having an application as possible, new, and sustainably sourced anti-tumor agent. In an Escherichia coli strain, optimized for sesquiterpene production, titers of 225 mg l−1 and 395 mg l−1, respectively, could be achieved. Remarkably, both enzymes share the same product profile thereby representing the first two terpene synthases from Basidiomycota with identical product profiles. We solved the crystal structure of Copu9 in its closed conformation, for the first time providing molecular details of sesquiterpene synthase from Basidiomycota. Based on the Copu9 structure, we conducted structure-based mutagenesis of amino acid residues lining the active site, thereby altering the product profile. Interestingly, the mutagenesis study also revealed that despite the conserved product profiles of Copu5 and Copu9 different conformational changes may accompany the catalytic cycle of the two enzymes. This observation suggests that the involvement of tertiary structure elements in the reaction mechanism(s) employed by terpene synthases may be more complex than commonly expected. Conclusion The presented product selectivity and titers of Copu5 and Copu9 may pave the way towards a sustainable, biotechnological production of the potentially new bioactive (+)-δ-cadinol. Furthermore, Copu5 and Copu9 may serve as model systems for further mechanistic studies of terpenoid catalysis. Graphical Abstract

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Section: Methodsmentioning

confidence: 99%

Biotechnological potential and initial characterization of two novel sesquiterpene synthases from Basidiomycota Coniophora puteana for heterologous production of δ-cadinol

et al. 2022

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“…50 µL of pyridine were added to each sample. Thereafter, 50 µL of MSTFA (N-Methyl-N-(trimethylsilyl)trifluoroacetamide) with 1% TCMS (Trimethylchlorosilane) was added and the samples were incubated in a water bath at 50°C for 1 h. GC-MS analysis was carried out using a modified version of a previously described protocol ( Ringel et al, 2020 ). The samples were analyzed using a Trace GC-MS Ultra system with DSQII (Thermo Scientific, USA).…”

Section: Methodsmentioning

confidence: 99%

Isolation and Investigation of Natural Rare Earth Metal Chelating Agents From Calothrix brevissima - A Step Towards Unraveling the Mechanisms of Metal Biosorption

Jurkowski

Paper

Brück

2022

Front. Bioeng. Biotechnol.

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In this study water soluble compounds that form complexes with Rare Earth Elements (REE) and other metals were isolated from Calothrix brevissima biomass with chromatographic methods for the first time. Molecular characterization showed that the isolated compounds are most likely polysaccharides comprised of arabinose, xylose, mannose, galactose and glucose. FT-IR analysis revealed functional groups involved in the binding mechanism of Tb are likely sulfate- and to a lesser extend hydroxyl-groups. The binding specificity of the isolated compounds was investigated with different metal solutions. Here, ions of the alkali and alkaline earth metals Na, K, Mg and Ca showed no competition for Tb-binding even at 10-fold excess concentration. Ions of the elements Co and Pb on the other hand replaced Tb at higher concentrations. Addition of the isolated compounds significantly reduced the precipitation of Eu at pH-values between 6.7 and 9.5, indicating that the interaction between the isolated chelators and Rare Earth Metals is stable even at high pH-values.

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“…Interestingly, the elisabethatriene diterpene macrocycle and its isoforms can act as biosynthetic precursors of the bioactive compounds erogorgiaene and pseudopterosin, having antibiotic and anti-inflammatory activities, respectively [4,5]. Unexpectedly, a single active site mutation, M75L, significantly shifts the product distribution and IE A becomes the dominant product (44%) in this enzyme variant [6].…”

Section: Introductionmentioning

confidence: 98%

“…Oxidation of IEs A and B by lipases results in the formation of the advanced pseudopterosin (P) precursor erogorgiaene and (1R)-epoxyelisabetha-5,14-diene (EED), respectively [6,7]. Ps, marine amphilectane-type diterpenoids from the gorgonian coral Antillogorgia elisabethae, feature superior anti-inflammatory properties which render them innovative target compounds for drug development [8,9].…”

Section: Introductionmentioning

confidence: 99%

Understanding the competing pathways leading to hydropyrene and isoelisabethatriene

Zev¹,

Ringel²,

Driller³

et al. 2022

Beilstein J. Org. Chem.

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Terpene synthases are responsible for the biosynthesis of terpenes, the largest family of natural products. Hydropyrene synthase generates hydropyrene and hydropyrenol as its main products along with two byproducts, isoelisabethatrienes A and B. Fascinatingly, a single active site mutation (M75L) diverts the product distribution towards isoelisabethatrienes A and B. In the current work, we study the competing pathways leading to these products using quantum chemical calculations in the gas phase. We show that there is a great thermodynamic preference for hydropyrene and hydropyrenol formation, and hence most likely in the synthesis of the isoelisabethatriene products kinetic control is at play.

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Towards a sustainable generation of pseudopterosin-type bioactives

Abstract: Pseudopterosins (Ps), marine diterpene glycosides derived from the marine octocoral Antillogorgia elisabethae, have potent anti-inflammatory activity demonstrated in phase II clinical trials. As multi-step total chemical synthesis is not economical,...

Cited by 10 publications

References 78 publications

Biotechnological potential and initial characterization of two novel sesquiterpene synthases from Basidiomycota Coniophora puteana for heterologous production of δ-cadinol

Biotechnological potential and initial characterization of two novel sesquiterpene synthases from Basidiomycota Coniophora puteana for heterologous production of δ-cadinol

Isolation and Investigation of Natural Rare Earth Metal Chelating Agents From Calothrix brevissima - A Step Towards Unraveling the Mechanisms of Metal Biosorption

Understanding the competing pathways leading to hydropyrene and isoelisabethatriene

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