Expression and Purification of Goat Lactoferrin from Pichia pastoris Expression System
| Autor: | I-Hua Chiang, Li-Jung Yin, Shann-Tzong Jiang, Gen-Hung Chen |
|---|---|
| Rok vydání: | 2007 |
| Předmět: |
DNA
Complementary Glycosylation Zeocin Iron Molecular Sequence Data Gene Expression Molecular cloning Chromatography Affinity Pichia Pichia pastoris law.invention chemistry.chemical_compound Affinity chromatography Sequence Analysis Protein law Animals Amino Acid Sequence Polyacrylamide gel electrophoresis biology Lactoferrin Goats biology.organism_classification Molecular biology Recombinant Proteins Yeast Biochemistry chemistry Recombinant DNA biology.protein Electrophoresis Polyacrylamide Gel Protein Binding Food Science |
| Zdroj: | Journal of Food Science. 72:M67-M71 |
| ISSN: | 1750-3841 0022-1147 |
| DOI: | 10.1111/j.1750-3841.2007.00281.x |
| Popis: | The recombinant goat lactoferrin (rGLF) was expressed in the methylotropic yeast Pichia pastoris using pGAPZalphaC vector, GAP as promoter, and Zeocin as the selective marker. After transformation of the GLF-pGAPZalphaC into Pichia pastoris X-33 expression host, the GLF-pGAPZalphaC vector was integrated into the GAP promoter locus of Pichia pastoris X-33 chromosome. The rGLF was expressed and secreted into the broth using alpha-factor preprosequence. SDS-PAGE and PAS staining analysis indicated that the rGLF could be purified to electrophoretic homogeneity by heparin-Sepharose 6 Fast Flow affinity chromatography and glycosylated by the expression host. The yield of purified rGLF was approximately 2.0 mg/L of culture broth. The N-terminal sequence was identical to the native goat lactoferrin (nGLF). The iron-binding behavior, papain-inhibiting property, and thermal stability of the purified rGLF were comparable to nGLF. This is the 1st report of intact goat lactoferrin expression using the P. pastoris system. |
| Databáze: | OpenAIRE |
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