Improved Production of Recombinant Myrosinase in <i>Pichia pastoris</i>
The effect of the deletion of a 57 bp native signal sequence, which transports the nascent protein through the endoplasmic reticulum membrane in plants, on improved <i>At</i>TGG1 plant myrosinase production in <i>Pichia pastoris</i> was studied. Myrosinase was extracellularly...
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| Auteurs principaux: | , , , , |
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| Format: | article |
| Langue: | EN |
| Publié: |
MDPI AG
2021
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| Sujets: | |
| Accès en ligne: | https://doaj.org/article/f9a3e6dad4c4487384b229e48f5920b5 |
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| Résumé: | The effect of the deletion of a 57 bp native signal sequence, which transports the nascent protein through the endoplasmic reticulum membrane in plants, on improved <i>At</i>TGG1 plant myrosinase production in <i>Pichia pastoris</i> was studied. Myrosinase was extracellularly produced in a 3-liter laboratory fermenter using α-mating factor as the secretion signal. After the deletion of the native signal sequence, both the specific productivity (164.8 U/L/h) and volumetric activity (27 U/mL) increased more than 40-fold compared to the expression of myrosinase containing its native signal sequence in combination with α-mating factor. The deletion of the native signal sequence resulted in slight changes in myrosinase properties: the optimum pH shifted from 6.5 to 7.0 and the maximal activating concentration of ascorbic acid increased from 1 mM to 1.5 mM. Kinetic parameters toward sinigrin were determined: 0.249 mM (K<sub>m</sub>) and 435.7 U/mg (V<sub>max</sub>). These results could be applied to the expression of other plant enzymes. |
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