Open Access

Multiple gene copy number increases total protein expression and enzyme activity of DNA topoisomerase I in Pichia pastoris

  • Authors:
    • Nur Adila Fadzil
    • Shern Kwok Lim
    • Ai Lan Chew
    • Boon Yin Khoo
  • View Affiliations

  • Published online on: August 9, 2022     https://doi.org/10.3892/wasj.2022.167
  • Article Number: 32
  • Copyright: © Fadzil et al. This is an open access article distributed under the terms of Creative Commons Attribution License.

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Abstract

The present study aimed to employ an in vivo strategy for the establishment of multi‑copy gene constructs of human DNA topoisomerase I (hTopI) using the pPIC3.5K vector in Pichia. The clones with multi‑copy inserts (His+ transformants) that were able to survive in the highest concentration of Geneticin® were found to express the highest expression level of total protein and exhibited the highest target enzyme activity. The highest level of total protein found was 1.76 mg/ml in GS115‑pPIC3.5K‑hTop1, which was resistant to 1.00 mg/ml Geneticin at 48 h of incubation. The highest enzyme activity of hTopI was also observed in the culture expressed by GS115‑pPIC3.5K‑hTopI, which was resistant to 1.00 mg/ml Geneticin® (19.7x104 Ul/OD600). On the whole, the present study provides information regarding the production of target protein from recombinant Pichia using only a shaker flask system, which can be further developed as an in‑house resource for screening potential anticancer agents.

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September-October 2022
Volume 4 Issue 5

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Spandidos Publications style
Fadzil NA, Lim SK, Chew AL and Khoo BY: Multiple gene copy number increases total protein expression and enzyme activity of DNA topoisomerase I in <em>Pichia pastoris</em>. World Acad Sci J 4: 32, 2022
APA
Fadzil, N.A., Lim, S.K., Chew, A.L., & Khoo, B.Y. (2022). Multiple gene copy number increases total protein expression and enzyme activity of DNA topoisomerase I in <em>Pichia pastoris</em>. World Academy of Sciences Journal, 4, 32. https://doi.org/10.3892/wasj.2022.167
MLA
Fadzil, N. A., Lim, S. K., Chew, A. L., Khoo, B. Y."Multiple gene copy number increases total protein expression and enzyme activity of DNA topoisomerase I in <em>Pichia pastoris</em>". World Academy of Sciences Journal 4.5 (2022): 32.
Chicago
Fadzil, N. A., Lim, S. K., Chew, A. L., Khoo, B. Y."Multiple gene copy number increases total protein expression and enzyme activity of DNA topoisomerase I in <em>Pichia pastoris</em>". World Academy of Sciences Journal 4, no. 5 (2022): 32. https://doi.org/10.3892/wasj.2022.167