Efficient evaluation of cellulose digestibility by Trichoderma reesei Rut-C30 cultures in online monitored shake flasks

Antonov, Elena; Wirth, Steffen; Gerlach, Tim; Schlembach, Ivan; Regestein, Lars

doi:10.1186/s12934-016-0567-7

Cited by 23 publications

(22 citation statements)

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“…During cultivation, the overall oxygen consumption (OT) can be determined by integrating the OTR. Antonov et al [46] recently analyzed the digestibility of different types of cellulose by online monitoring of the respiration activity of T. reesei Rut-C30. Alternative cellulase producers were subsequently investigated using this methodology, demonstrating its high potential [47].…”

Section: Introductionmentioning

confidence: 99%

Online evaluation of the metabolic activity of Ustilago maydis on (poly)galacturonic acid

et al. 2018

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BackgroundPectin is a rather complex and highly branched polysaccharide strengthening the plant cell wall. Thus, many different pectinases are required for an efficient microbial conversion of biomass waste streams with a high pectin content like citrus peel, apple pomace or sugar beet pulp. The screening and optimization of strains growing on pectic substrates requires both, quantification of the residual substrate and an accurate determination of the enzymatic activity. Galacturonic acid, the main sugar unit of pectin, is an uncommon substrate for microbial fermentations. Thus, growth and enzyme production of the applied strain has to be characterized in detail to understand the microbial system. An essential step to reach this goal is the development of online monitoring tools.ResultsIn this study, a method for the online determination of residual substrate was developed for the growth of the plant pathogenic fungus Ustilago maydis on pectic substrates such as galacturonic acid. To this end, an U. maydis strain was used that expressed a heterologous exo-polygalacturonase for growth on polygalacturonic acid. The growth behavior on galacturonic acid was analyzed by online measurement of the respiration activity. A method for the online prediction of the residual galacturonic acid concentration during the cultivation, based on the overall oxygen consumption, was developed and verified by offline sampling. This sensitive method was extended towards polygalacturonic acid, which is challenging to quantify via offline measurements. Finally, the enzymatic activity in the culture supernatant was calculated and the enzyme stability during the course of the cultivation was confirmed.ConclusionThe introduced method can reliably predict the residual (poly)galacturonic acid concentration based on the overall oxygen consumption. Based on this method, the enzymatic activity of the culture broth of an U. maydis strain expressing a heterologous exo-polygalacturonase could be calculated. It was demonstrated that the method is especially advantageous for determination of low enzymatic activities. In future, it will be applied to U. maydis strains in which the number of produced hydrolytic enzymes is increased for more efficient degradation.Electronic supplementary materialThe online version of this article (10.1186/s13036-018-0128-1) contains supplementary material, which is available to authorized users.

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Section: Introductionmentioning

confidence: 99%

Online evaluation of the metabolic activity of Ustilago maydis on (poly)galacturonic acid

et al. 2018

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“…An important factor affecting the product price is the cost of the sugar feedstock (9,10). Agricultural crops such as sugarcane, sugar beet, or corn are the main sources for sugar feedstock, and the utilization of cellulosic biomass containing abundant units of glucose is also intensively investigated (11)(12)(13). However, due to the competition between food production and sugar feedstock production from agricultural crops and the current high costs of the pretreatment and conversion processes of utilizing cellulosic biomass (14), novel sustainable and economical sources for sugar feedstock and related conversion technologies should be explored.…”

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confidence: 99%

Effects of Reduced and Enhanced Glycogen Pools on Salt-Induced Sucrose Production in a Sucrose-Secreting Strain of Synechococcus elongatus PCC 7942

Qiao

Duan

Zhang

et al. 2018

Appl Environ Microbiol

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Sucrose and glycogen syntheses in cyanobacteria share the common precursor glucose-1-phosphate. It is generally assumed that lowering glycogen synthesis could drive more carbon toward sucrose synthesis that can be induced by salt stress among cyanobacteria. By using a theophylline-dependent riboswitch system, the expression of , a key gene in glycogen synthesis, was downregulated in a quantitative manner in a sucrose-secreting strain of PCC 7942. We observed that the stepwise suppression of glycogen synthesis limited rather than stimulated sucrose production in the salt-stressed cells, suggesting that glycogen could serve as a carbon pool for the synthesis of sucrose. Accordingly, we generated glycogen-overproducing strains, but the increased glycogen pool alone did not stimulate sucrose production, indicating that alternative steps limit the carbon flux toward the synthesis of sucrose. Consistent with previous studies that showed that sucrose-phosphate synthase (SPS) catalyzes the rate-limiting step in sucrose synthesis, the combination of glycogen overproduction and overexpression resulted in increased sucrose production. Our results indicate that the glycogen and sucrose pools are closely linked in PCC 7942, and we propose that enhancing the glycogen pool could be a promising strategy for the improvement of sucrose production by cyanobacteria in the presence of a strong sucrose synthesis sink. Many cyanobacteria naturally synthesize and accumulate sucrose when stressed by NaCl, which provides novel possibilities for obtaining sugar feedstock by engineering of cyanobacteria. It has been assumed that glycogen synthesis competes with sucrose synthesis for the carbon flux. However, our results showed that the suppression of glycogen synthesis decreased rather than stimulated sucrose production in a sucrose-secreting strain of PCC 7942. This result suggests that glycogen could serve as a supportive rather than a competitive carbon pool for the synthesis of sucrose, providing new insights about the relation between glycogen synthesis and sucrose synthesis in cyanobacteria. This finding is also useful to guide metabolic engineering work to optimize the production of sucrose and possibly other products by cyanobacteria.

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“…These values corresponded to 18 and 5 FPU/g cellulose , respectively, and are close to the cellulase loading of 10 FPU/g cellulose , typically used for separate cellulose hydrolysis (43). Since cellulose digestibility has a major impact on cellulose hydrolysis, 120 g/L of highly digestible Sigmacell as well as 120 g/L recalcitrant α-cellulose were tested as substrates (44). All media were buffered to pH 6.5 using 100 mM MES buffer.…”

Section: Introductionmentioning

confidence: 65%

Consolidated Bioprocessing of Cellulose to Itaconic Acid by a Co-Culture of Trichoderma Reesei and Ustilago Maydis.

Schlembach

Tehrani

Blank

et al. 2020

Preprint

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Background: Itaconic acid is a bio-derived platform chemical with uses ranging from polymer synthesis to biofuel production. The efficient conversion of cellulosic waste streams into itaconic acid could thus enable the sustainable production of a variety of substitutes for fossil oil based products. However, the realization of such a process is currently hindered by an expensive conversion of cellulose into fermentable sugars. Here, we present the stepwise development of a fully consolidated bioprocess (CBP), which is capable to directly convert recalcitrant cellulose into itaconic acid without the need for separate cellulose hydrolysis including the application of commercial cellulases. The process is based on a synthetic microbial consortium of the cellulase producer Trichoderma reesei and the itaconic acid producing yeast Ustilago maydis. Results: The efficiency of the process was compared to a simultaneous hydrolysis and fermentation setup (SSF). Because of the additional substrate consumption of T. reesei in the CBP, the itaconic acid yield was significantly lower in the CBP than in the SSF. In order to increase yield and productivity of itaconic acid in the CBP, the population dynamics was manipulated by varying the inoculation delay between T. reesei and U. maydis. Surprisingly, neither inoculation delay nor inoculation density significantly affected the population development or the CBP performance. Instead, the substrate availability was the most important parameter. U. maydis was only able to grow and to produce itaconic acid when the cellulose concentration and thus, the sugar supply rate, was high. Finally, the metabolic processes during fed-batch CBP were analyzed in depth by online respiration measurements. A method was developed to estimate cellulose consumption, itaconic acid formation as well as the actual itaconic acid production yield online. Again, substrate availability was the key factor also controlling itaconic acid yield. In summary, an itaconic acid titer of 34 g/L with a total productivity of up to 0.07 g/L/h and a yield of 0.16 g/g could be reached during fed-batch cultivation. Conclusion: This study demonstrates the feasibility of consortium based CBP for itaconic acid production and also lays the fundament for the development and improvement of similar microbial consortia for cellulose based organic acid production.

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Efficient evaluation of cellulose digestibility by Trichoderma reesei Rut-C30 cultures in online monitored shake flasks

Cited by 23 publications

References 65 publications

Online evaluation of the metabolic activity of Ustilago maydis on (poly)galacturonic acid

Online evaluation of the metabolic activity of Ustilago maydis on (poly)galacturonic acid

Effects of Reduced and Enhanced Glycogen Pools on Salt-Induced Sucrose Production in a Sucrose-Secreting Strain of Synechococcus elongatus PCC 7942

Consolidated Bioprocessing of Cellulose to Itaconic Acid by a Co-Culture of Trichoderma Reesei and Ustilago Maydis.

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