Artículo
Pichia pastoris engineering for the production of a modified phospholipase C
Elena, Claudia Edicht
; Ravasi, Pablo
; Cerminati, Sebastián
; Peirú, Salvador
; Castelli, Maria Eugenia
; Menzella, Hugo Gabriel
Fecha de publicación:
12/2016
Editorial:
Elsevier
Revista:
Process Biochemistry
ISSN:
1359-5113
Idioma:
Inglés
Tipo de recurso:
Artículo publicado
Clasificación temática:
Resumen
Crude vegetable oils are refined to remove impurities that adversely impact in their stability, color and flavor. In recent years, enzymatic degumming methods using phospholipase C (PLC) enzymes provide an environmentally friendly process for phospholipids removal with improved oil recovery yields. In this study, Pichia pastoris was used as the expression system for the production of PLC-Y, a modified PLC enzyme derived from Bacillus cereus. Production of secreted PLC-Y driven by the methanol inducible AOX1 promoter was optimized by genetic strain engineering which included gene codon optimization, generation of multi-copy chromosomal integrations and the co-expression of helper factors supporting protein folding, processing and secretion processes. In addition, tunable promoters directing helper factor expression were tested. In batch cultures, a strain harboring seven integrated copies of the PLC-Y expression cassette and co-expressing the HAC1 transcription factor under an attenuated AOX1 promoter showed a 6.2 fold increase in the production titers compared to the strain harboring a single gene copy. A fed-batch fermentation process developed using this engineered strain produced 4.5 g/l of this enzyme. The results presented in this work show the viability of using PLC-Y for oil degumming and provide a manufacturing process for its cost effective production.
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Articulos(IPROBYQ)
Articulos de INST. DE PROCESOS BIOTECNOLOGICOS Y QUIMICOS ROSARIO
Articulos de INST. DE PROCESOS BIOTECNOLOGICOS Y QUIMICOS ROSARIO
Citación
Elena, Claudia Edicht; Ravasi, Pablo; Cerminati, Sebastián; Peirú, Salvador; Castelli, Maria Eugenia; et al.; Pichia pastoris engineering for the production of a modified phospholipase C; Elsevier; Process Biochemistry; 51; 12; 12-2016; 1935-1944
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