Optimization of conditions for secretion of dengue virus type 2 envelope domain III using Pichia pastoris

被引：28

作者：

Batra, Gaurav ^{[1
]}

Gurramkonda, Chandrasekhar ^{[1
]}

Nemani, Satish Kumar ^{[1
]}

Jain, Swatantra Kumar ^{[2
]}

Swaminathan, Sathyamangalam ^{[1
]}

Khanna, Navin ^{[1
]}

机构：

[1] Int Ctr Genet Engn & Biotechnol, RGP Grp, New Delhi 110067, India

[2] Jamia Hamdard, Dept Biotechnol, New Delhi 110062, India

来源：

JOURNAL OF BIOSCIENCE AND BIOENGINEERING | 2010年 / 110卷 / 04期

关键词：

Dengue; Flavivirus; Envelope domain III; Pichia pastoris; Fed-batch cultivation; Vaccine; NEUTRALIZING ANTIBODIES; ESCHERICHIA-COLI; PROTEIN; EXPRESSION; GLYCOPROTEIN; INDUCTION; EPITOPES; ANTIGEN; VACCINE; BINDING;

D O I：

10.1016/j.jbiosc.2010.05.001

中图分类号：

Q81 [生物工程学（生物技术）]; Q93 [微生物学];

学科分类号：

071005 ; 0836 ; 090102 ; 100705 ;

摘要：

We have developed a recombinant clone of the methylotrophic yeast Pichia pastoris capable of secreting dengue virus type 2 envelope domain III (sEDIII-2). We explored various induction parameters including media composition, temperature, pH, and methanol concentration, to optimize conditions for sEDIII-2 expression in shake flask culture. Induction at 20 degrees C in the presence of 2% (v/v) methanol in a medium buffered to pH 5.8 resulted in highest secretion of 5EDIII-2. This yield could be further enhanced up to 70% by repeated induction of the same initial biomass. Using a fed-batch cultivation strategy, we observed that shake-flask yields can be scaled up similar to 8-fold in a bioreactor. We obtained similar to 94% purity with >70% recovery after purification. This study, which demonstrates for the first time the feasibility of secreting envelope domain III using the P. pastoris host, will be relevant to sub-unit approaches to dengue vaccine development. (C) 2010, The Society for Biotechnology, Japan. All rights reserved.

引用

页码：408 / 414

页数：7

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