標題: Expression, characterization, and purification of recombinant porcine lactoferrin in Pichia pastoris
作者: Wang, SH
Yang, TS
Lin, SM
Tsai, MS
Wu, SC
Mao, SJT
生物科技學系
Department of Biological Science and Technology
公開日期: 1-六月-2002
摘要: Recombinant porcine lactoferrin (rPLF) was synthesized in Pichia pastoris using a constitutive promoter from the glyceraldehyde-3-phosphate dehydrogenase gene. Strains expressing rPLF with its own signal sequence or with that from the yeast alpha-mating factor (alpha-MF) were able to produce and secrete rPLF, but levels were consistently higher using alpha-MF constructs. In contrast, P. pastoris strains that expressed rPLF without a signal sequence produced the protein in an insoluble intracellular form. Increasing the initial pH of shake-flask culture medium from 6.0 to 7.0 or adding ferric ions to the medium (to 100 muM) resulted irk significant improvements in expression of rPLF from P. pastoris. Expression levels (approximately 12 mg/L) were much higher than those observed from Saccharomyces cerevisiae strains (1-2 mg/L). P pastoris-secreted rPLF was isolated and purified via a one-step simple procedure using a heparin column. The molecular size (78 kDa), isoelectric point (8.8-9.0), N-terminal amino acid sequence, and iron-binding capability of rPLF were each similar to that of native milk PLF. (C) 2002 Elsevier Science (USA).
URI: http://dx.doi.org/10.1006/prep.2001.1607
http://hdl.handle.net/11536/28757
ISSN: 1046-5928
DOI: 10.1006/prep.2001.1607
期刊: PROTEIN EXPRESSION AND PURIFICATION
Volume: 25
Issue: 1
起始頁: 41
結束頁: 49
顯示於類別:期刊論文


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