Molecular gene cloning and overexpression of the phytase from Debaryomyces castellii CBS 2923
| Autor: | Mélanie Ragon, Patrick Chemardin, Virginie Neugnot-Roux, Hélène Boze, G. Moulin |
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| Přispěvatelé: | Ingénierie des Réactions Biologiques, Bio-productions (IR2B), Université Montpellier 2 - Sciences et Techniques (UM2)-Institut National de la Recherche Agronomique (INRA), Adisseo France SAS |
| Jazyk: | angličtina |
| Rok vydání: | 2008 |
| Předmět: |
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Debaryomyces castellii Gene isolation Molecular Sequence Data Molecular cloning Pichia Pichia pastoris [SDV.BBM]Life Sciences [q-bio]/Biochemistry Molecular Biology Amino Acid Sequence Cloning Molecular Gene Phylogeny 6-Phytase biology Base Sequence biology.organism_classification Phytase Molecular biology Yeast Recombinant Proteins Alcohol oxidase Kinetics Biochemistry Saccharomycetales Heterologous expression Biotechnology |
| Zdroj: | Protein Expression and Purification Protein Expression and Purification, Elsevier, 2008, 58 (2), pp.275-283. ⟨10.1016/j.pep.2007.12.003⟩ |
| ISSN: | 1046-5928 1096-0279 |
| Popis: | The ORF encoding the Debaryomyces castellii CBS 2923 phytase was isolated. The deduced 461-amino-acid sequence corresponded to a 51.2 kDa protein and contained the consensus motif (RHGXRXP) which is conserved among phytases. No signal sequence cleavage site was detected. Nine potential N -glycosylation sites have been predicted. The protein shared 21–69% sequence identities with various phytases of yeast or fungal origin. Heterologous expression of the D. castellii CBS 2923 phytase in the methylotrophic yeast Pichia pastoris was tested under both the P. pastoris inducible alcohol oxidase (AOX1) promoter and the constitutive glyceraldehyde-3-phosphate dehydrogenase (GAP) promoter. Maximum production levels obtained were 476 U ml −1 , with the AOX1 expression system and 16.5 U ml −1 with the GAP one. These productions corresponded to a 320-fold and a 10-fold overexpression of the protein, respectively as compared to the homologous production. The biochemical characteristics of the recombinant phytase were identical to those of the native enzyme. |
| Databáze: | OpenAIRE |
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