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The growth of Pichia pastoris Mut+ on methanol–glycerol mixtures fits to interactive dual-limited kinetics: model development and application to optimised fed-batch operation for heterologous protein production

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Abstract

The methanol–glycerol co-feeding during the induction stage for heterologous protein production in Pichia pastoris has shown significant productive applications. Available model analysis applied to this dual-limited condition is scarce and normally does not consider the interaction effects between the substrates. In this work, a dual-limited growth model of P. pastoris considering an interactive kinetic effect was applied to an optimised fed-batch process production of heterologous Rhizopus oryzae lipase (ROL). In the proposed model, the growth kinetics on glycerol is fully expressed, whereas methanol kinetics is modulated by the co-metabolisation of glycerol, resulting in an enhancing effect of glycerol-specific growth rate. The modelling approach of fed-batch cultures also included the methanol volatilisation caused by the aeration that was found to be a not-negligible phenomenon. The model predicts the ability of P. pastoris to keep control of the methanol concentration in the broth during ROL-optimised production process in fed batch and fits satisfactorily the specific cell growth rate and ROL production. Implications of interaction effect are discussed applying the general procedure of modelling approach.

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Abbreviations

µ:

Specific growth rate (h−1)

µin :

Enhancing growth rate parameter (h−1)

D:

Dilution rate (h−1)

X:

Biomass concentration (g l−1)

V:

Volume of broth (l)

S:

Substrate concentration (g l−1)

KS :

Half-saturation constant (g l−1)

KI :

Methanol inhibition constant (g l−1)

q:

Specific substrate consumption rate (g gX−1 h−1)

YX/G :

Glycerol yield coefficient (gX gG−1)

YX/M :

Methanol yield coefficient (gX gM−1)

\({Y_{X/G}}^{{Ov}}\) :

Overall glycerol yield coefficient (gX gG−1)

\({Y_{X/M}}^{{Ov}}\) :

Overall methanol yield coefficient (gX gM−1)

kev :

Volatilisation first-order constant (h−1)

qP :

Specific ROL productivity (UA gX−1 h−1)

F:

Feed rate (l h−1)

β(0, 1, 2, 3, 4, 5) :

Polynomial coefficients

Max:

Maximal

0:

Initial

M:

Methanol

G:

Glycerol

1:

Metabolic glycerol pathway

2:

Metabolic methanol pathway

f:

Feed

Des:

Designed

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Acknowledgements

This work was supported by Project N°11110486 from Fondo Nacional de Ciencia y Tecnología (FONDECYT, Chile). C. Canales wishes to thank Comisión Nacional de Investigación Científica y Tecnológica (CONICYT, Chile) for scholarship support.

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Authors and Affiliations

  1. Escuela de Ingeniería Bioquímica, Pontificia Universidad Católica de Valparaíso, Av. Brasil 2085, Valparaiso, 2340000, Chile

    Christian Canales, Claudia Altamirano & Julio Berrios

  2. Facultad de Ingeniería y Tecnología, Universidad San Sebastián, Lientur 1457, Concepción, 4080871, Chile

    Christian Canales

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  1. Christian Canales
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Correspondence to Julio Berrios.

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Canales, C., Altamirano, C. & Berrios, J. The growth of Pichia pastoris Mut+ on methanol–glycerol mixtures fits to interactive dual-limited kinetics: model development and application to optimised fed-batch operation for heterologous protein production. Bioprocess Biosyst Eng 41, 1827–1838 (2018). https://doi.org/10.1007/s00449-018-2005-1

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