Developmental studies of <i>Claviceps paspali</i> seed cultures for the submerged production of lysergic acid derivatives

Sočić, H.; Gaberc-Porekar, Vladka; Pertot, E.; Puc, A.; Miličić, S.

doi:10.1002/jobm.3620260906

Cited by 5 publications

(1 citation statement)

References 17 publications

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“…C. paspali has long been used in the pharmaceutical industry to produce lysergic acid and similar ergot alkaloids in submerged liquid cultures (Ricicová et al 1982; Socic et al 1986). Nevertheless, the targeted molecular genetic modification of this fungus remained a major challenge due to the low efficiencies of most steps in the transformation workflow, including protoplast regeneration, DNA introduction, mitotically stable integration of the incoming DNA, and homogenization of the transformant allele.…”

Section: Discussionmentioning

confidence: 99%

Inactivation of the indole-diterpene biosynthetic gene cluster of Claviceps paspali by Agrobacterium-mediated gene replacement

Kozák

Szilágyi

Vágó

et al. 2018

Appl Microbiol Biotechnol

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The hypocrealean fungus Claviceps paspali is a parasite of wild grasses. This fungus is widely utilized in the pharmaceutical industry for the manufacture of ergot alkaloids, but also produces tremorgenic and neurotoxic indole-diterpene (IDT) secondary metabolites such as paspalitrems A and B. IDTs cause significant losses in agriculture and represent health hazards that threaten food security. Conversely, IDTs may also be utilized as lead compounds for pharmaceutical drug discovery. Current protoplast-mediated transformation protocols of C. paspali are inadequate as they suffer from inefficiencies in protoplast regeneration, a low frequency of DNA integration, and a low mitotic stability of the nascent transformants. We adapted and optimized Agrobacterium tumefaciens-mediated transformation (ATMT) for C. paspali and validated this method with the straightforward creation of a mutant strain of this fungus featuring a targeted replacement of key genes in the putative IDT biosynthetic gene cluster. Complete abrogation of IDT production in isolates of the mutant strain proved the predicted involvement of the target genes in the biosynthesis of IDTs. The mutant isolates continued to produce ergot alkaloids undisturbed, indicating that equivalent mutants generated in industrial ergot producers may have a better safety profile as they are devoid of IDT-type mycotoxins. Meanwhile, ATMT optimized for Claviceps spp. may open the door for the facile genetic engineering of these industrially and ecologically important organisms.

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Section: Discussionmentioning

confidence: 99%

Inactivation of the indole-diterpene biosynthetic gene cluster of Claviceps paspali by Agrobacterium-mediated gene replacement

Kozák

Szilágyi

Vágó

et al. 2018

Appl Microbiol Biotechnol

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Physiological control and process kinetics of clavine alkaloid production by Claviceps purpurea

Flieger¹,

Votruba²,

K?en³

et al. 1988

Appl Microbiol Biotechnol

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Summary. Kinetic parameters of production of clavine alkaloids were evaluated in two Claviceps purpurea strains. Mutagenesis brought about enhanced resistance of the biosynthetic system towards alkaloids. Addition of glucose into the fermentation medium altered the zero order kinetics of production to activation-inhibition kinetics. The glucose treatment allowed performance of both "elymoclavine-inhibitionless" and "clavine alkaloid-decompositionless" fermentations if a combination of fermentation and separation units in a closed loop was used. Nomenlacture :kl, rate constant of agroclavine synthesis (mg Agro. mg Elymo/1 -g DW. day for stage 1, mg Agro/g DW. day for stage 2); k2, parameter describing inhibition of agroclavine formation rate by elymoclavine (mg Elymo/1); k3, specific rate of agroclavine decay (1/g DW-day); k4, maximal specific rate of elymoclavine synthesis (stage 1, 1/g DW. day, stage 2, mg Elymo/g DW-day); kz, maximal specific rate of elymoclavine synthesis in stage 1 (inhibition-activation mechanism) (mg Elymo/g DW. day); ks, physiological constant describing the elymoclavine decay rate (12/g DW. day-mg Elymo); ks, physiological constant describing the activation of elymoclavine biosynthesis by elymoclavine (mg Elymo/1); k6, physiological constant describing the repression of elymoclavine biosynthesis by elymoclavine (rag Elymo/1); k7, maximal specific growth rate (1/day); ks, specific rate of biomass decay (1/g DW. day); A, agroclavine concentration (rag/l); E, elymoclavine concentration (mg/l); rA, specific rate of agroclavine biosynthesis (mg Agro/g DW-day); rE, specific rate of elymoclavine biosynthesis (mg Elymo/g DW. day); ri, specific rate of alkaloid biosynthesis (mg alkaloid/g DW. day); X, dry biomass concentration (g/l);/l, specific growth rate (1/day) Offprint requests to: M. Flieger

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Physiological control and process kinetics of clavine alkaloid production byClaviceps purpurea

Flieger

Votruba

Křen

et al. 1988

Appl Microbiol Biotechnol

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Developmental studies of Claviceps paspali seed cultures for the submerged production of lysergic acid derivatives

Cited by 5 publications

References 17 publications

Inactivation of the indole-diterpene biosynthetic gene cluster of Claviceps paspali by Agrobacterium-mediated gene replacement

Inactivation of the indole-diterpene biosynthetic gene cluster of Claviceps paspali by Agrobacterium-mediated gene replacement

Physiological control and process kinetics of clavine alkaloid production by Claviceps purpurea

Physiological control and process kinetics of clavine alkaloid production byClaviceps purpurea

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