Development of chemically defined media to express trp-analog-labeled proteins in a lactococcus lactis trp auxotroph

Development of chemically defined media to express trp-analog-labeled proteins in a lactococcus lactis trp auxotroph

Author Shao, Jinfeng Google Scholar
Marcondes, Marcelo F. M. Autor UNIFESP Google Scholar
Oliveira, Vitor Autor UNIFESP Google Scholar
Broos, Jaap Google Scholar
Abstract Chemically defined media for growth of Lactococcus lactis strains contain about 50 components, making them laborious and expensive growth media. However, they are crucial for metabolism studies as well as for expression of heterologous proteins labeled with unnatural amino acids. In particular, the L. lactis Trp auxotroph PA1002, over-expressing the tryptophanyl tRNA synthetase enzyme of L. lactis, is very suitable for the biosynthetic incorporation of Trp analogs in proteins because of its most relaxed substrate specificity reported towards Trp analogs. Here we present two much simpler defined media for L. lactis, which consist of only 24 or 31 components, respectively, and with which the L. lactis Trp auxotroph shows similar growth characteristics as with a 50-component chemically defined medium. Importantly, the expression levels of two recombinant proteins used for evaluation were up to 2-3 times higher in these new media than in the 50-component medium, without affecting the Trp analog incorporation efficiency. Taken together, the simplest chemically defined media reported so far for L. lactis are presented. Since L. lactis also shows auxotrophy for Arg, His, Ile, Leu Val, and Met, our simplified media may also be useful for the biosynthetic incorporation of analogs of these five amino acids. (C) 2016 The Author(s) Published by S. Karger AG, Basel
Keywords Alloprotein
Lactococcus Lactis
Synthetic Medium
Tryptophan AnalogTryptophan Analogs
Biosynthetic Incorporation
Fluorescence Spectroscopy
Streptococcus-Cremoris
Growth
Overproduction
Requirements
Host
Language English
Sponsor China Scholarship Council
FAPESP
Grant number This work was financially supported by the China Scholarship Council and FAPESP. We thank Dr. B.W. Dijkstra for critical reading of the manuscript.
Date 2016
Published in Journal Of Molecular Microbiology And Biotechnology. Basel, v. 26, n. 4, p. 269-276, 2016.
ISSN 1464-1801 (Sherpa/Romeo, impact factor)
Publisher Pontificia Universidade Catolica Campinas
Extent 269-276
Origin https://doi.org/10.1159/000445687
Access rights Open access Open Access
Type Article
Web of Science ID WOS:000381109500004
URI http://repositorio.unifesp.br/handle/11600/49447

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