The control of promoter activity by oxygen availability appears to be an intriguing system for heterologous protein production. In fact, during cell growth in a bioreactor, an oxygen shortage is easily obtained simply by interrupting the air supply. The purpose of our work was to explore the possible use of hypoxic induction of the KIPDC1 promoter to direct heterologous gene expression in yeast. In the present study, an expression system based on the KIPDC1 promoter was developed and characterized. Several heterologous proteins, differing in size, origin, localization, and posttranslational modification, were successfully expressed in Kluyveromyces lactis under the control of the wild type or a modified promoter sequence, with a production ratio between 4 and more than 100. Yields were further optimized by a more accurate control of hypoxic physiological conditions. Production of as high as 180 mg/liter of human interleukin-1β was obtained, representing the highest value obtained with yeasts in a lab-scale bioreactor to date. Moreover, the transferability of our system to related yeasts was assessed. The lacZ gene from Escherichia coli was cloned downstream of the KIPDC1 promoter in order to get β-galactosidase activity in response to induction of the promoter. A centromeric vector harboring this expression cassette was introduced in Saccharomyces cerevisiae and in Zygosaccharomyces bailii, and effects of hypoxic induction were measured and compared to those already observed in K. lactis cells. Interestingly, we found that the induction still worked in Z. bailii; thus, this promoter constitutes a possible inducible system for this new nonconventional host. Copyright © 2007, American Society for Microbiology. All Rights Reserved.
Camattari, A., Bianchi, M.M., Branduardi, P., Porro, D., & Brambilla, L.G. (2007). Induction by hypoxia of heterologous proteins production with the KlPDC1 promoter in yeasts. APPLIED AND ENVIRONMENTAL MICROBIOLOGY, 73(3), 922-929 [10.1128/AEM.01764-06].
|Citazione:||Camattari, A., Bianchi, M.M., Branduardi, P., Porro, D., & Brambilla, L.G. (2007). Induction by hypoxia of heterologous proteins production with the KlPDC1 promoter in yeasts. APPLIED AND ENVIRONMENTAL MICROBIOLOGY, 73(3), 922-929 [10.1128/AEM.01764-06].|
|Tipo:||Articolo in rivista - Articolo scientifico|
|Carattere della pubblicazione:||Scientifica|
|Titolo:||Induction by hypoxia of heterologous proteins production with the KlPDC1 promoter in yeasts.|
|Autori:||Camattari, A; Bianchi, MM; Branduardi, P; Porro,D; Brambilla, LG|
|Data di pubblicazione:||2007|
|Rivista:||APPLIED AND ENVIRONMENTAL MICROBIOLOGY|
|Digital Object Identifier (DOI):||http://dx.doi.org/10.1128/AEM.01764-06|
|Appare nelle tipologie:||01 - Articolo su rivista|