Effect of temperature on recombinant protein production using the Bm5/Bm5.NPV expression system

Authors

  • J. N. Andersen,

    1. Pharmaceutical Production Research Facility, Faculty of Engineering, The University of Calgary, Calgary, AB, Canada T2N 1N4
    Current affiliation:
    1. Chemical Engineering Dept., State University of New York, 908 Furnas Hall, Buffalo, NY 14260–4200, U.S.A
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  • P. G. Sriram,

    1. Pharmaceutical Production Research Facility, Faculty of Engineering, The University of Calgary, Calgary, AB, Canada T2N 1N4
    Current affiliation:
    1. Chemical Engineering Dept., State University of New York, 908 Furnas Hall, Buffalo, NY 14260–4200, U.S.A
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  • N. Kalogerakis,

    Corresponding author
    1. Pharmaceutical Production Research Facility, Faculty of Engineering, The University of Calgary, Calgary, AB, Canada T2N 1N4
    Current affiliation:
    1. Chemical Engineering Dept., State University of New York, 908 Furnas Hall, Buffalo, NY 14260–4200, U.S.A
    • Pharmaceutical Production Research Facility, Faculty of Engineering, The University of Calgary, Calgary, AB, Canada T2N 1N4
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  • L. A. Behie

    1. Pharmaceutical Production Research Facility, Faculty of Engineering, The University of Calgary, Calgary, AB, Canada T2N 1N4
    Current affiliation:
    1. Chemical Engineering Dept., State University of New York, 908 Furnas Hall, Buffalo, NY 14260–4200, U.S.A
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Abstract

A series of experiments have been conducted using a recombinant baculovirus/insect cell expression system (Bm5/Bm5.NPV.CAT) to establish the optimum temperature for both cell growth and virus infection. Bm5 cell growth was found to be limited at temperatures below 22°C and ceased completely at temperatures above 34°C. In the range between 24 and 28°C, final cell densities always reached 96% of the highest achievable viable cell density. The shortest population doubling time was obtained at 28°C. Overall, a consistent increase in metabolism with increasing temperatures was observed. During the infection/viral replication phase, an increase in the temperature from 25 to 31°C resulted in a faster decrease in viable cell density and an earlier production of chloramphenicol acetyltransferase (CAT). Furthermore, protein yield at temperatures above 28°C was significantly reduced. Overall, the best temperature for the infection phase for the Bm5/Bm5.NPV expression system was found to be 25°C when the cells are cultured in serum free media.

Abstract

On a mené une série d'expériences à l'aide d'un système d'expression de cellules d'insectes et de baculovirus recombinant (Bm5Bm5.NPV/P5) afin d'établir la température optimale pour la croissance des cellules et l'infection par le virus. On a trouvé que la croissance des cellules Bm5 était limitée aux températures inférieures à 22°C et qu'elle cessait complètement aux températures supérieures à 34°C. Entre 24 et 28°C, les densités de cellules finales atteignent toujours 96% de la plus haute densité de cellules viables qui puissent ětre atteintes. Le temps le plus court de doublement de la population a été obtenu à 28°C. Globalement, une augmentation consistante du métabolisme a été observé avec l'augmentation des températures. Lors de la phase de replication virale/infection, une augmentation de la température de 25 à 31°C a entraǐné une diminution plus rapide de la densité de cellules viables et une production plus précoce de chloramphénicol acétyltransférase (CAT). En outre, le rendement en protéines à des températures supérieures à 28°C est réduit de manière significative. De façon générale, on a trouvé que la meilleure température pour la phase d'infection pour le système d'expression Bm5/Bm5.NPV était de 25°C lorsque les cellules sont cultivées dans un milieu sans sérum.

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