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OPTIMISATION OF PROTEIN
EXPRESSION AND SECRETION IN LACTOBACILLI
Main goal:
Developing genetic tools for controlled and optimised production
of proteins and peptides in selected Lactobacillus species.
Sub-goals:
1. Characterise regulated, strong promoters from Lactobacillus,
with emphasis on promoters from bacteriocin operons.
2. Construction of vectors for optimisation of expression. These
studies will involve attempts to develop fully food-grade expression
systems. The systems will be evaluated in fermentor studies.
3. Integrate pertinent gene constructs into the genome, to obtain
stable heritage and to avoid undesired spreading of genes.
4. To study, compare and evaluate different secretion machineries
(sec-dependent and sec-independent).
5. To over-express industrially commercially interesting heterologous
proteins and peptides, for example bacteriocins, eukaryotic antimicrobial
peptides (AMPs; e.g lactoferricin, AMP from fish), chitinases, amylases
and peptidases.
6. Examine the constructed strains in food fermentations. In this
way one can study the significance of single genes in situ during
food production.
Summary:
Certain species of lactobacilli constitute a highly attractive group
of bacteria, mainly because their extended use in food applications.
The use of these bacteria as production organisms is generally regarded
as less effective than the use of aerobic organisms such as E. coli
or Bacillus species. An enormous potential, however, lies in the
possibility for in situ production of technological or therapeutic
proteins in fermented food (and feed). The potential has not been
fully exploited because of lack of knowledge concerning gene expression
and protein secretion in lactobacilli. Using their very strong position
in Lactobacillus research as a starting point, the applicants aim
at developing tools that permit (regulated) over-expression and
secretion of proteins/peptides at industrially interesting levels.
The continuous accumulation of knowledge about the genetic systems
(regulated and constitutive strong promoters, different types of
secretion machinery) will be accompanied by evaluation and optimisation
in fermentor studies and in in situ fermentations. The work will
focus on bacteriocins from lactobacilli as a follow-up on previous,
partly co-operative research at all three laboratories, but other
proteins such as chitinases, amylases and peptidases will be studied.
A considerable part of the to-be-developed technology is already
protected by patents held by the partners.
Partners:
NLH (I.F. Nes, V.G.H. Eijsink, G. Mathiesen, J. Blatny)
MATFORSK (L. Axelsson, K. Naterstad, E. Sørvig)
SINTEF-Kjemi, Trondheim (I.M. Aasen)
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Lokalkontakt på PEP: Geir Mathiesen
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