Scalable methanol-free production of recombinant glucuronoyl esterase in Pichia pastoris.
BMC Res Notes
; 12(1): 596, 2019 Sep 18.
Article
em En
| MEDLINE
| ID: mdl-31533815
ABSTRACT
OBJECTIVE:
Glucuronoyl esterase (GE) is an emerging enzyme that improves fractionation of lignin-carbohydrate complexes. However, the commercial availability of GE is limited, which hinders the research of GE-based bioprocesses for its industrial application in lignocellulose biorefineries. This study evaluated a workable, cost-effective, and commercially scalable production strategy to improve the ease of GE-based research. This strategy consisted of a constitutive and methanol-free enzyme production step coupled with a two-step filtration process. The aim was to determine if this strategy can yield copious amounts of GE, by secretion into the extracellular medium with an acceptable purity that could allow its direct application. This approach was further validated for cellobiose dehydrogenase, another emerging lignocellulose degrading enzyme which is scarcely available at high cost.RESULTS:
The secreted recombinant enzymes were functionally produced in excess of levels previously reported for constitutive production (1489-2780 mg L-1), and were secreted at moderate to high percentages of the total extracellular protein (51-94%). The constant glycerol feed, implemented during fed-batch fermentation, lead to a decline in growth rate and plateaued productivity. Tangential flow ultrafiltration was used to concentrate cell-free enzyme extracts 5-6-fold, reaching enzyme activity levels (1020-202 U L-1) that could allow their direct application.Palavras-chave
Texto completo:
1
Coleções:
01-internacional
Base de dados:
MEDLINE
Assunto principal:
Proteínas Recombinantes
/
Ácido Glucurônico
/
Esterases
Idioma:
En
Revista:
BMC Res Notes
Ano de publicação:
2019
Tipo de documento:
Article
País de afiliação:
África do Sul