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World Journal of Microbiology and Biotechnology

, Volume 29, Issue 8, pp 1421–1429 | Cite as

Scale-up from shake flasks to bioreactor, based on power input and Streptomyces lividans morphology, for the production of recombinant APA (45/47 kDa protein) from Mycobacterium tuberculosis

  • Ramsés A. Gamboa-Suasnavart
  • Luz D. Marín-Palacio
  • José A. Martínez-Sotelo
  • Clara Espitia
  • Luis Servín-González
  • Norma A. Valdez-Cruz
  • Mauricio A. Trujillo-Roldán
Original Paper

Abstract

Culture conditions in shake flasks affect filamentous Streptomyces lividans morphology, as well the productivity and O-mannosylation of recombinant Ala-Pro-rich O-glycoprotein (known as the 45/47 kDa or APA antigen) from Mycobacterium tuberculosis. In order to scale up from previous reported shake flasks to bioreactor, data from the literature on the effect of agitation on morphology of Streptomyces strains were used to obtain gassed volumetric power input values that can be used to obtain a morphology of S. lividans in bioreactor similar to the morphology previously reported in coiled/baffled shake flasks by our group. Morphology of S. lividans was successfully scaled-up, obtaining similar mycelial sizes in both scales with diameters of 0.21 ± 0.09 mm in baffled and coiled shake flasks, and 0.15 ± 0.01 mm in the bioreactor. Moreover, the specific growth rate was successfully scaled up (0.09 ± 0.02 and 0.12 ± 0.01 h−1, for bioreactors and flasks, respectively), and the recombinant protein productivity measured by densitometry, as well. More interestingly, the quality of the recombinant glycoprotein measured as the amount of mannoses attached to the C-terminal of APA was also scaled- up; with up to five mannose residues in cultures carried out in shake flasks; and six in the bioreactor. However, final biomass concentration was not similar, indicating that although the process can be scaled-up using the power input, others factors like oxygen transfer rate, tip speed or energy dissipation/circulation function can be an influence on bacterial metabolism.

Keywords

Scale-up Power input APA 45/47 kDa O-mannosylation Mycobacteriumtuberculosis Streptomyceslividans 

Abbreviations

Di

Impeller diameter (m)

Fg

Volumetric gas flow rate (m3/min)

g

Gravitational acceleration (g/m2)

N

Agitation speed (rev/min)

Np

Power number (Dimensionless)

P

Power input by agitation (W)

Pg

Gassed power input (W)

Pg*

Corrected gassed power input by bioreactor geometry (W)

Re

Reynolds number (Dimensionless)

V

Operation volume (m3)

W

Impeller blade width (m)

µ

Fluid viscosity (mPa.s)

ρ

Density (kg/m3)

Notes

Acknowledgments

This work was financed by CONACYT-INNOVAPYME 181895, CONACYT 178528, 104951-Z and PAPPIT-UNAM IN-209113, IN-210013. RGS thanks CONACyT scholarship (316929). Authors thank Celia Flores, M. Sc., and Enrique Galindo, PhD. (Instituto de Biotecnología, UNAM), and Marisol Córdova, PhD. (Centro de Ciencias Aplicadas y Desarrollo Tecnológico, UNAM) for their technical assistance in image analysis. Authors thank Erendida Garcia, Chem. (Instituto de Química, UNAM) for MALDI-TOF analysis. We also thank Ana Delgado for reviewing the English version of the manuscript.

Conflict of interest

The authors declare that they have no conflict of interest.

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Copyright information

© Springer Science+Business Media Dordrecht 2013

Authors and Affiliations

  • Ramsés A. Gamboa-Suasnavart
    • 1
    • 2
  • Luz D. Marín-Palacio
    • 1
    • 2
  • José A. Martínez-Sotelo
    • 3
  • Clara Espitia
    • 3
  • Luis Servín-González
    • 2
  • Norma A. Valdez-Cruz
    • 2
  • Mauricio A. Trujillo-Roldán
    • 1
    • 2
  1. 1.Unidad de Bioprocesos, Instituto de Investigaciones BiomédicasUniversidad Nacional Autónoma de MéxicoMexicoMexico
  2. 2.Departamento de Biología Molecular y Biotecnología, Instituto de Investigaciones BiomédicasUniversidad Nacional Autónoma de MéxicoMexicoMexico
  3. 3.Departamento de Inmunología, Instituto de Investigaciones BiomédicasUniversidad Nacional Autónoma de MéxicoMexicoMexico

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