請用此 Handle URI 來引用此文件: http://hdl.handle.net/11455/38265
標題: Production of recombinant porcine lactoferrin exhibiting antibacterial activity in methylotrophic yeast, Pichia pastoris
作者: 陳全木
Chen, H.L.
Lai, Y.W.
Yen, C.C.
Lin, Y.Y.
Lu, C.Y.
Yang, S.H.
Tsai, T.C.
Lin, Y.J.
Lin, C.W.
Chen, C.M.
關鍵字: recombinant lactoferrin
bactericidal peptide
methylotrophic yeast
culture
Pichia pastoris
bovine lactoferrin
binding protein
growth-hormone
expression
peptides
milk
purification
secretion
membrane
lysozyme
期刊/報告no:: Journal of Molecular Microbiology and Biotechnology, Volume 8, Issue 3, Page(s) 141-149.
摘要: Lactoferrin is a metal-binding glycoprotein exhibiting multifunctional immunoregulation of antibacterial, antioxidant, anti-endotoxin and antiviral activities. Uptake of porcine lactoferrin (PLF) has been shown to enhance resistance to diarrhea and anemia in neonatal piglets. In this study, the methylotrophic yeast, Pichia pastoris, was used to express a recombinant PLF (rPLF) gene from swine mammary gland. A synthetic secretion cassette was constructed using the inducible promoter of the alcohol oxidase-1 gene (AOX1) and the yeast a-mating factor signal peptide. After electroporation and Zeocin selection, several clones expressed high levels of rPLF protein which constitutes more than 30% of the total protein. A time-course study showed that rPLF mRNA transcripts are stably expressed during 120 h of culture induction. rPLF was exported into the culture supernatant at approximately 87 mg/l and a large portion of rPLF was accumulated in the cell cytoplasm at approximately 760 mg/l after 72 h of methanol induction. Recombinant PLF protein was purified via a heparin column using a fast protein liquid chromatography system. The glycosylation of P. pastoris-derived rPLF was analyzed and similar patterns to milk PLF were observed. Pepsin hydrolysate of rPLF displayed high bactericidal activity against Escherichia coli ATCC 25922 under scanning electron microscopy observation and minimal inhibitory concentration and minimal bactericidal concentration tests. Our results suggested that the methylotrophic yeast-inducible system is suitable for large-scale production of active antibacterial rPLF glycoprotein. Copyright (C) 2004 S. Karger AG, Basel.
URI: http://hdl.handle.net/11455/38265
ISSN: 1464-1801
文章連結: http://dx.doi.org/10.1159/000085787
顯示於類別:生命科學系所

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