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ACS Synth Biol. 2018 Feb 16;7(2):452-461. doi: 10.1021/acssynbio.7b00285. Epub 2018 Jan 19.

Sporobeads: The Utilization of the Bacillus subtilis Endospore Crust as a Protein Display Platform.

Author information

1
Institute of Microbiology, Technische Universität (TU) Dresden , 01062 Dresden, Germany.

Abstract

Upon starvation, the soil bacterium Bacillus subtilis forms an intracellular, metabolically inactive endospore. Its core contains the DNA, encased by three protein layers protecting it against a multitude of environmental threats. The outermost layer, the crust, harbors great potential as a protein-displaying platform: a gene of interest can be translationally fused to a crust protein gene, resulting in endospores displaying the desired protein on their surface. To unlock this potential in a standardized fashion, we designed a suite of 12 vectors (Sporovectors), based on the BioBrick cloning standard. With these vectors, proteins can easily be fused N- or C-terminally to the six crust proteins CotV, CotW, CotX, CotY, CotZ, and CgeA under the control of the strongest crust gene promoter PcotYZ. All Sporovectors were evaluated with GFP and two different laccases. On the basis of our data, CotY and CotZ represent the best anchor proteins. But there are significant differences in activity and functional stability between the two tested laccases. Our vector suite is a powerful tool to generate and evaluate a vast variety of functionalized endospores. It allows quickly identifying the best anchor and fusion site for the protein of interest. Our findings demonstrate that the crust of B. subtilis endospores is an inexpensive and easy platform for displaying different proteins of interest.

KEYWORDS:

Bacillus vectors; BioBrick standard; protein immobilization; sporulation; surface display

PMID:
29284082
DOI:
10.1021/acssynbio.7b00285
[Indexed for MEDLINE]

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