Stable Multicopy Vectors for High–Level Secretion of Recombinant Human Serum Albumin by Kluyveromyces Yeasts
Autor: | Gerard Jung, N. Amellal, Patrice Yeh, Isabelle Maury, Reinhard Fleer, Jerome Becquart, Jean-François Mayaux, F. Bacchetta, P. Baduel, Alain Fournier, H. L'Hôte, H. Fukuhara |
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Rok vydání: | 1991 |
Předmět: |
Genetic Vectors
Molecular Sequence Data Restriction Mapping Serum albumin Gene Expression Gene dosage law.invention Kluyveromyces law Gene expression medicine Humans Secretion Codon Promoter Regions Genetic Serum Albumin Expression vector Base Sequence biology Human serum albumin biology.organism_classification Molecular biology Recombinant Proteins Biochemistry Fermentation Recombinant DNA biology.protein Biotechnology medicine.drug |
Zdroj: | Bio/Technology. 9:968-975 |
ISSN: | 0733-222X |
DOI: | 10.1038/nbt1091-968 |
Popis: | We have designed stable pKD1 derivatives for efficient secretion of recombinant human serum albumin (rHSA) by industrial strains of Kluyveromyces yeasts. A comparison of this multi-copy expression system with isogenic cassettes integrated at chromosomal loci demonstrated that high level secretion of rHSA is a function of gene dosage in K. lactis. Various signal sequences could be used, and the secretion levels were independent of the presence of the native pro peptide. The mitotic stability of the pKD1-based expression vectors was found to be species and strain dependent and was influenced by promoter strength and culture conditions. Vector stability was drastically enhanced when the HSA gene was expressed from an inducible promoter: 90% of the transformed cells still harbored the vector after 100 generations of non-selective growth in uninduced culture conditions. Secretion levels in the range of several grams per liter of correctly folded and processed rHSA were obtained at the pilot scale, thus making the industrial production of pharmaceutical-grade, Kluyveromyces-derived rHSA economically feasible. |
Databáze: | OpenAIRE |
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