Dataset Information

Membrane modulation of super-secreting “midiBacillus” expressing the major Staphylococcus aureus antigen – a mass-spectrometry based absolute quantification approach.

ABSTRACT: Bacillus subtilis is a Gram-positive bacterium considered as a “cell-factory” for industrial enzymes and biopharmaceuticals. Given the commercial advantage of this organism, researchers have been working towards the improvement of this organism as a producer of secreted proteins. Nonetheless, there is the need to obtain more knowledge regarding the absolute composition of the gram positive cell membrane in order to increase the secretion capability of B. subtilis and gain insight into potential bottlenecks in protein production and/or secretion. However, membrane proteins are one of the most challenging classes of proteins, mainly due to their high hydrophobicity and low abundance compared to their soluble counterparts. To this regard, we established a method for global characterization of absolute protein abundances within the membrane of B. subtilis and verified the effects of large scale protein export on a super-secreting strain. We did shotgun proteomics on three different fractions (extracellular, cytosol and membrane) and spiked in UPS2 proteins in each of these to calculate absolute protein abundances. The absolute values obtained from the shotgun experiment were validated by targeted proteomics. Furthermore, we performed western blot analysis on the three subcellular fractions using antibodies against proteins localized in these three different compartments in order to visually validate the enrichment of membrane proteins in the corresponding fraction.

INSTRUMENT(S): TSQ Vantage, LTQ Orbitrap Elite

ORGANISM(S): Bacillus Subtilis Subsp. Subtilis

SUBMITTER: Minia Antelo

LAB HEAD: Dörte Becher

PROVIDER: PXD015496 | Pride | 2020-03-20

REPOSITORIES: Pride

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Publications

Membrane Modulation of Super-Secreting "midiBacillus" Expressing the Major Staphylococcus aureus Antigen - A Mass-Spectrometry-Based Absolute Quantification Approach.

Antelo-Varela Minia M Aguilar Suárez Rocío R Bartel Jürgen J Bernal-Cabas Margarita M Stobernack Tim T Sura Thomas T van Dijl Jan Maarten JM Maaß Sandra S Becher Dörte D

Frontiers in bioengineering and biotechnology 20200228

Bacillus subtilis has been extensively used as a microbial cell factory for industrial enzymes due to its excellent capacities for protein secretion and large-scale fermentation. This bacterium is also an attractive host for biopharmaceutical production. However, the secretion potential of this organism is not fully utilized yet, mostly due to a limited understanding of critical rearrangements in the membrane proteome upon high-level protein secretion. Recently, it was shown that bottlene ...[more]

PMID: 32185169

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Dataset Information

Membrane modulation of super-secreting “midiBacillus” expressing the major Staphylococcus aureus antigen – a mass-spectrometry based absolute quantification approach.

Publications

Membrane Modulation of Super-Secreting "midi<i>Bacillus</i>" Expressing the Major <i>Staphylococcus aureus</i> Antigen - A Mass-Spectrometry-Based Absolute Quantification Approach.

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