Premium
Optimization of fed‐batch production of the model recombinant protein GFP in Lactococcus lactis
Author(s) -
Oddone Gian M.,
Lan Christopher Q.,
Rawsthorne Helen,
Mills David A.,
Block David E.
Publication year - 2006
Publication title -
biotechnology and bioengineering
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.136
H-Index - 189
eISSN - 1097-0290
pISSN - 0006-3592
DOI - 10.1002/bit.21192
Subject(s) - lactococcus lactis , recombinant dna , green fluorescent protein , fermentation , bacteria , hemin , nisin , chemistry , food science , biochemistry , biology , lactic acid , chromatography , enzyme , heme , genetics , gene
Optimization of recombinant protein production using lactic acid bacteria (LAB) remains an important obstacle on the road to realizing LAB as oral vaccine delivery vehicles. Despite this, there have been few published investigations to explore the higher limits of LAB recombinant protein expression in fed‐batch fermentations. In this study, results from response surface experiments suggested an optimal set of conditions for expression of green fluorescent protein (GFP), a model recombinant protein, in bench‐scale, fed‐batch Lactococcus lactis IL1403 fermentations. The 48 4‐L fed‐batch fermentations in this set of experiments, along with preliminary studies, investigated the effects of pH, temperature, hemin concentration, concentration of the nisin inducer per cell, and time of induction. Cell densities in this data set ranged from 2.9 to 7.4 g/L and maximum GFP expression per cell ranged from 0.1 to 4.4 relative fluorescence units (RFU)/g. The optimal 4‐L, fed‐batch fermentation process found here yields growth and protein expression values that dramatically improve upon results from traditional test tube and flask processes. Relative to the traditional process, the experimental optimum conditions yield 4.9 times the cell density, 1.6 times the protein per cell mass, and 8 times the total protein concentration. Unexpectedly, experiments also revealed that the compound hemin, known previously to improve growth and survival of Lactococcus lactis ( L. lactis ), negatively impacted recombinant protein production when added in concentrations from 5 to 20 µg/mL with this strain. The improvement in protein expression over traditional processes demonstrated here is an important step toward commercial development of LAB for oral delivery of recombinant vaccines and therapeutic proteins. Biotechnol. Bioeng. 2007;96:1127–1138. © 2006 Wiley Periodicals, Inc.