Mervi Toivari, Maija-Leena Vehkomäki, Laura Ruohonen, Merja Penttilä, Marilyn G. Wiebe
{"title":"利用磷酸葡萄糖异构酶缺陷的酿酒酵母生产 d-葡萄糖酸","authors":"Mervi Toivari, Maija-Leena Vehkomäki, Laura Ruohonen, Merja Penttilä, Marilyn G. Wiebe","doi":"10.1007/s10529-023-03443-2","DOIUrl":null,"url":null,"abstract":"<p><span>d</span>-Glucaric acid is a potential biobased platform chemical. Previously mainly <i>Escherichia coli,</i> but also the yeast <i>Saccharomyces cerevisiae,</i> and <i>Pichia pastoris,</i> have been engineered for conversion of <span>d</span>-glucose to <span>d</span>-glucaric acid via myo-inositol. One reason for low yields from the yeast strains is the strong flux towards glycolysis. Thus, to decrease the flux of <span>d</span>-glucose to biomass, and to increase <span>d</span>-glucaric acid yield, the four step <span>d</span>-glucaric acid pathway was introduced into a phosphoglucose isomerase deficient (Pgi1p-deficient) <i>Saccharomyces cerevisiae</i> strain. High <span>d</span>-glucose concentrations are toxic to the Pgi1p-deficient strains, so various feeding strategies and use of polymeric substrates were studied. Uniformly labelled <sup>13</sup>C-glucose confirmed conversion of <span>d</span>-glucose to <span>d</span>-glucaric acid. In batch bioreactor cultures with pulsed <span>d</span>-fructose and ethanol provision 1.3 g <span>d</span>-glucaric acid L<sup>−1</sup> was produced. The <span>d</span>-glucaric acid titer (0.71 g <span>d</span>-glucaric acid L<sup>−1</sup>) was lower in nitrogen limited conditions, but the yield, 0.23 g <span>d</span>-glucaric acid [g <span>d</span>-glucose consumed]<sup>−1</sup>, was among the highest that has so far been reported from yeast. Accumulation of myo-inositol indicated that myo-inositol oxygenase activity was limiting, and that there would be potential to even higher yield. The Pgi1p-deficiency in <i>S. cerevisiae</i> provides an approach that in combination with other reported modifications and bioprocess strategies would promote the development of high yield <span>d</span>-glucaric acid yeast strains.</p>","PeriodicalId":8929,"journal":{"name":"Biotechnology Letters","volume":null,"pages":null},"PeriodicalIF":2.0000,"publicationDate":"2023-12-08","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":"0","resultStr":"{\"title\":\"Production of d-glucaric acid with phosphoglucose isomerase-deficient Saccharomyces cerevisiae\",\"authors\":\"Mervi Toivari, Maija-Leena Vehkomäki, Laura Ruohonen, Merja Penttilä, Marilyn G. Wiebe\",\"doi\":\"10.1007/s10529-023-03443-2\",\"DOIUrl\":null,\"url\":null,\"abstract\":\"<p><span>d</span>-Glucaric acid is a potential biobased platform chemical. Previously mainly <i>Escherichia coli,</i> but also the yeast <i>Saccharomyces cerevisiae,</i> and <i>Pichia pastoris,</i> have been engineered for conversion of <span>d</span>-glucose to <span>d</span>-glucaric acid via myo-inositol. One reason for low yields from the yeast strains is the strong flux towards glycolysis. Thus, to decrease the flux of <span>d</span>-glucose to biomass, and to increase <span>d</span>-glucaric acid yield, the four step <span>d</span>-glucaric acid pathway was introduced into a phosphoglucose isomerase deficient (Pgi1p-deficient) <i>Saccharomyces cerevisiae</i> strain. High <span>d</span>-glucose concentrations are toxic to the Pgi1p-deficient strains, so various feeding strategies and use of polymeric substrates were studied. Uniformly labelled <sup>13</sup>C-glucose confirmed conversion of <span>d</span>-glucose to <span>d</span>-glucaric acid. In batch bioreactor cultures with pulsed <span>d</span>-fructose and ethanol provision 1.3 g <span>d</span>-glucaric acid L<sup>−1</sup> was produced. The <span>d</span>-glucaric acid titer (0.71 g <span>d</span>-glucaric acid L<sup>−1</sup>) was lower in nitrogen limited conditions, but the yield, 0.23 g <span>d</span>-glucaric acid [g <span>d</span>-glucose consumed]<sup>−1</sup>, was among the highest that has so far been reported from yeast. Accumulation of myo-inositol indicated that myo-inositol oxygenase activity was limiting, and that there would be potential to even higher yield. 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Production of d-glucaric acid with phosphoglucose isomerase-deficient Saccharomyces cerevisiae
d-Glucaric acid is a potential biobased platform chemical. Previously mainly Escherichia coli, but also the yeast Saccharomyces cerevisiae, and Pichia pastoris, have been engineered for conversion of d-glucose to d-glucaric acid via myo-inositol. One reason for low yields from the yeast strains is the strong flux towards glycolysis. Thus, to decrease the flux of d-glucose to biomass, and to increase d-glucaric acid yield, the four step d-glucaric acid pathway was introduced into a phosphoglucose isomerase deficient (Pgi1p-deficient) Saccharomyces cerevisiae strain. High d-glucose concentrations are toxic to the Pgi1p-deficient strains, so various feeding strategies and use of polymeric substrates were studied. Uniformly labelled 13C-glucose confirmed conversion of d-glucose to d-glucaric acid. In batch bioreactor cultures with pulsed d-fructose and ethanol provision 1.3 g d-glucaric acid L−1 was produced. The d-glucaric acid titer (0.71 g d-glucaric acid L−1) was lower in nitrogen limited conditions, but the yield, 0.23 g d-glucaric acid [g d-glucose consumed]−1, was among the highest that has so far been reported from yeast. Accumulation of myo-inositol indicated that myo-inositol oxygenase activity was limiting, and that there would be potential to even higher yield. The Pgi1p-deficiency in S. cerevisiae provides an approach that in combination with other reported modifications and bioprocess strategies would promote the development of high yield d-glucaric acid yeast strains.
期刊介绍：
Biotechnology Letters is the world’s leading rapid-publication primary journal dedicated to biotechnology as a whole – that is to topics relating to actual or potential applications of biological reactions affected by microbial, plant or animal cells and biocatalysts derived from them.
All relevant aspects of molecular biology, genetics and cell biochemistry, of process and reactor design, of pre- and post-treatment steps, and of manufacturing or service operations are therefore included.
Contributions from industrial and academic laboratories are equally welcome. We also welcome contributions covering biotechnological aspects of regenerative medicine and biomaterials and also cancer biotechnology. Criteria for the acceptance of papers relate to our aim of publishing useful and informative results that will be of value to other workers in related fields.
The emphasis is very much on novelty and immediacy in order to justify rapid publication of authors’ results. It should be noted, however, that we do not normally publish papers (but this is not absolute) that deal with unidentified consortia of microorganisms (e.g. as in activated sludge) as these results may not be easily reproducible in other laboratories.
Papers describing the isolation and identification of microorganisms are not regarded as appropriate but such information can be appended as supporting information to a paper. Papers dealing with simple process development are usually considered to lack sufficient novelty or interest to warrant publication.
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