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Production, secretion and purification of a correctly folded staphylococcal antigen in Lactococcus lactis

Overview of attention for article published in Microbial Cell Factories, July 2015
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  • Above-average Attention Score compared to outputs of the same age (55th percentile)
  • Above-average Attention Score compared to outputs of the same age and source (64th percentile)

Mentioned by

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1 tweeter
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1 Facebook page
googleplus
1 Google+ user

Citations

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12 Dimensions

Readers on

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34 Mendeley
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Title
Production, secretion and purification of a correctly folded staphylococcal antigen in Lactococcus lactis
Published in
Microbial Cell Factories, July 2015
DOI 10.1186/s12934-015-0271-z
Pubmed ID
Authors

Frédéric Samazan, Bachra Rokbi, Delphine Seguin, Fabienne Telles, Valérie Gautier, Gilbert Richarme, Didier Chevret, Paloma Fernández Varela, Christophe Velours, Isabelle Poquet

Abstract

Lactococcus lactis, a lactic acid bacterium traditionally used to ferment milk and manufacture cheeses, is also, in the biotechnology field, an interesting host to produce proteins of medical interest, as it is "Generally Recognized As Safe". Furthermore, as L. lactis naturally secretes only one major endogenous protein (Usp45), the secretion of heterologous proteins in this species facilitates their purification from a protein-poor culture medium. Here, we developed and optimized protein production and secretion in L. lactis to obtain proteins of high quality, both correctly folded and pure to a high extent. As proteins to be produced, we chose the two transmembrane members of the HtrA protease family in Staphylococcus aureus, an important extra-cellular pathogen, as these putative surface-exposed antigens could constitute good targets for vaccine development. A recombinant ORF encoding a C-terminal, soluble, proteolytically inactive and tagged form of each staphylococcal HtrA protein was cloned into a lactococcal expression-secretion vector. After growth and induction of recombinant gene expression, L. lactis was able to produce and secrete each recombinant rHtrA protein as a stable form that accumulated in the culture medium in similar amounts as the naturally secreted endogenous protein, Usp45. L. lactis growth in fermenters, in particular in a rich optimized medium, led to higher yields for each rHtrA protein. Protein purification from the lactococcal culture medium was easily achieved in one step and allowed recovery of highly pure and stable proteins whose identity was confirmed by mass spectrometry. Although rHtrA proteins were monomeric, they displayed the same secondary structure content, thermal stability and chaperone activity as many other HtrA family members, indicating that they were correctly folded. rHtrA protein immunogenicity was established in mice. The raised polyclonal antibodies allowed studying the expression and subcellular localization of wild type proteins in S. aureus: although both proteins were expressed, only HtrA1 was found to be, as predicted, exposed at the staphylococcal cell surface suggesting that it could be a better candidate for vaccine development. In this study, an efficient process was developed to produce and secrete putative staphylococcal surface antigens in L. lactis and to purify them to homogeneity in one step from the culture supernatant. This allowed recovering fully folded, stable and pure proteins which constitute promising vaccine candidates to be tested for protection against staphylococcal infection. L. lactis thus proved to be an efficient and competitive cell factory to produce proteins of high quality for medical applications.

Twitter Demographics

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Mendeley readers

The data shown below were compiled from readership statistics for 34 Mendeley readers of this research output. Click here to see the associated Mendeley record.

Geographical breakdown

Country Count As %
Brazil 1 3%
Unknown 33 97%

Demographic breakdown

Readers by professional status Count As %
Researcher 7 21%
Student > Ph. D. Student 5 15%
Student > Bachelor 4 12%
Student > Master 4 12%
Librarian 1 3%
Other 2 6%
Unknown 11 32%
Readers by discipline Count As %
Agricultural and Biological Sciences 10 29%
Biochemistry, Genetics and Molecular Biology 5 15%
Immunology and Microbiology 2 6%
Chemical Engineering 1 3%
Computer Science 1 3%
Other 3 9%
Unknown 12 35%

Attention Score in Context

This research output has an Altmetric Attention Score of 2. This is our high-level measure of the quality and quantity of online attention that it has received. This Attention Score, as well as the ranking and number of research outputs shown below, was calculated when the research output was last mentioned on 16 August 2015.
All research outputs
#6,649,700
of 11,878,541 outputs
Outputs from Microbial Cell Factories
#410
of 861 outputs
Outputs of similar age
#102,422
of 238,016 outputs
Outputs of similar age from Microbial Cell Factories
#10
of 31 outputs
Altmetric has tracked 11,878,541 research outputs across all sources so far. This one is in the 43rd percentile – i.e., 43% of other outputs scored the same or lower than it.
So far Altmetric has tracked 861 research outputs from this source. They receive a mean Attention Score of 3.6. This one has gotten more attention than average, scoring higher than 51% of its peers.
Older research outputs will score higher simply because they've had more time to accumulate mentions. To account for age we can compare this Altmetric Attention Score to the 238,016 tracked outputs that were published within six weeks on either side of this one in any source. This one has gotten more attention than average, scoring higher than 55% of its contemporaries.
We're also able to compare this research output to 31 others from the same source and published within six weeks on either side of this one. This one has gotten more attention than average, scoring higher than 64% of its contemporaries.