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Expression and Purification of Goat Lactoferrin from Pichia pastoris Expression System
Author(s) -
Chen GenHung,
Yin LiJung,
Chiang IHua,
Jiang ShannTzong
Publication year - 2007
Publication title -
journal of food science
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.772
H-Index - 150
eISSN - 1750-3841
pISSN - 0022-1147
DOI - 10.1111/j.1750-3841.2007.00281.x
Subject(s) - pichia pastoris , lactoferrin , recombinant dna , pichia , biology , microbiology and biotechnology , expression vector , affinity chromatography , yeast , sepharose , biochemistry , gene , enzyme
The recombinant goat lactoferrin (rGLF) was expressed in the methylotropic yeast Pichia pastoris using pGAPZαC vector, GAP as promoter, and Zeocin as the selective marker. After transformation of the GLF‐pGAPZαC into Pichia pastoris X‐33 expression host, the GLF‐pGAPZαC vector was integrated into the GAP promoter locus of Pichia pastoris X‐33 chromosome. The rGLF was expressed and secreted into the broth using α‐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.