Expression and Antibacterial Activity of Recombinant Human Lactoferrin in Methylotrophic Yeast, Pichia pastoris

Methylotrophic Yeast, Pichia pastoris에서 사람 락토페린의 발현 및 항균성 연구

  • Lee Sang O (Department of Bioscience and Biotechnology, College of Natural Sciences, Hankook University of Foreign Studies) ;
  • Im Eun Mi (Department of Bioscience and Biotechnology, College of Natural Sciences, Hankook University of Foreign Studies) ;
  • Nam Eun Joo (Department of Bioscience and Biotechnology, College of Natural Sciences, Hankook University of Foreign Studies) ;
  • Lee Hyune Hwan (Department of Bioscience and Biotechnology, College of Natural Sciences, Hankook University of Foreign Studies)
  • 이상오 (한국외국어대학교 자연과학대학 생명공학과) ;
  • 임은미 (한국외국어대학교 자연과학대학 생명공학과) ;
  • 남은주 (한국외국어대학교 자연과학대학 생명공학과) ;
  • 이현환 (한국외국어대학교 자연과학대학 생명공학과)
  • Published : 2004.12.01

Abstract

The expression and antibacterial. activity of recombinant human lactoferrin (hLf) was studied from meth­ylotrophic yeast Pichia pastoris. The gene encoding hLf, isolated from human breast cDNA library, was subcloned into the expression vector, pPIC3.5K under the control of AOX1 promoter. The gene was integrated into the host chromosome and was identified by Southern blotting. The expression of the integrated gene was investigated by RT-PCR, Northern blotting, SDS-PAGE and Western blotting. Discrete band corresponding to hLf was detected from the SDS-PAGE, which was confirmed by Western blotting. The expression was also confirmed by RT-PCR and Northern blotting. The antibacterial activity of the recombinant hLf (rhLf) was investigated using Staphy­lococcus aureus ATCC 6538P and Micrococcus flavus ATCC 10240 as test organisms. The rhLf showed strong antibacterial activities against the bacteria. Furthermore, many Gram-negative animal pathogens such as E.coli ATCC8739, 25922, and Salmonella typhimurium 114 and 115, Pseudomonas fluorescens ID 963 I, P. aeruginosa KCCM 11802, and Gram-positive bacteria Bacillus mesentericus were also inhibited in their growth by the rhLf.

Keywords

antibacterial activity;expression;Pichia pastoris;recombinant human lactoferrin

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