Truncated prosequence of rhizopus oryzae lipase : key factor for production improvement and biocatalyst stability
López Fernández, Josu (Universitat Autònoma de Barcelona. Departament d'Enginyeria Química, Biològica i Ambiental)
Barrero Peña, Juan José (Universitat Autònoma de Barcelona. Departament d'Enginyeria Química, Biològica i Ambiental)
Benaiges, M. Dolors (Universitat Autònoma de Barcelona. Departament d'Enginyeria Química, Biològica i Ambiental)
Valero Barranco, Francisco (Universitat Autònoma de Barcelona. Departament d'Enginyeria Química, Biològica i Ambiental)

Data:	2019
Resum:	Recombinant Rhizopus oryzae lipase (mature sequence, rROL) was modified by adding to its N-terminal 28 additional amino acids from the C-terminal of the prosequence (proROL) to obtain a biocatalyst more suitable for the biodiesel industry. Both enzymes were expressed in Pichia pastoris and compared in terms of production bioprocess parameters, biochemical properties, and stability. Growth kinetics, production, and yields were better for proROL harboring strain than rROL one in batch cultures. When different fed-batch strategies were applied, lipase production and volumetric productivity of proROL-strain were always higher (5. 4 and 4. 4-fold, respectively) in the best case. rROL and proROL enzymatic activity was dependent on ionic strength and peaked in 200 mM Tris-HCl buffer. The optimum temperature and pH for rROL were influenced by ionic strength, but those for proROL were not. The presence of these amino acids altered lipase substrate specificity and increased proROL stability when different temperature, pH, and methanol/ethanol concentrations were employed. The 28 amino acids were found to be preferably removed by proteases, leading to the transformation of proROL into rROL. Nevertheless, the truncated prosequence enhanced Rhizopus oryzae lipase heterologous production and stability, making it more appropriate as industrial biocatalyst.
Ajuts:	Ministerio de Economía y Competitividad BES-2017-080858 Ministerio de Economía y Competitividad CTQ2016-74959-R Agència de Gestió d'Ajuts Universitaris i de Recerca 2017/SGR-1462
Drets:	Aquest document està subjecte a una llicència d'ús Creative Commons. Es permet la reproducció total o parcial, la distribució, la comunicació pública de l'obra i la creació d'obres derivades, fins i tot amb finalitats comercials, sempre i quan es reconegui l'autoria de l'obra original.
Llengua:	Anglès
Document:	Article ; recerca ; Versió publicada
Matèria:	Rhizopus oryzae lipase ; P. pastoris ; Biocatalysis ; Fed-batch cultures ; Prosequence ; Stability
Publicat a:	Catalysts, Vol. 9, issue 11 (Nov. 2019) , art. 961, ISSN 2073-4344

DOI: 10.3390/catal9110961

16 p, 2.6 MB

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