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A PopZ-Linked Apical Recruitment Assay for Studying Protein-Protein Interactions in the Bacterial Cell Envelope.

08:00 EDT 24th September 2019 | BioPortfolio

Summary of "A PopZ-Linked Apical Recruitment Assay for Studying Protein-Protein Interactions in the Bacterial Cell Envelope."

Most bacteria are surrounded by a complex cell envelope. As with many biological processes, studies of envelope assembly have benefited from cell-based assays for detecting protein-protein interactions. These assays use simple readouts and lack a protein purification requirement, making them ideal for early-stage investigations. The most widely used two-hybrid interaction assay for proteins involved in envelope biogenesis is based on the reconstitution of adenylate cyclase activity from a split enzyme. Because adenylate cyclase is only functional in the cytoplasm, both protein fusions used in the assay must have a terminus located in this compartment. However, many envelope assembly factors are wholly extracytoplasmic. Detecting interactions involving such proteins using two-hybrid systems has therefore been problematic. To address this issue, we developed a cytological assay in Escherichia coli based on PopZ from Caulobacter crescentus. Here, we demonstrate the utility of this PopZ-Linked Apical Recruitment (POLAR) method for detecting interactions between proteins located in different cellular compartments. Additionally, we report that recruitment of an active peptidoglycan synthase to the cell pole is detrimental for E. coli and that interactions between proteins in the inner and outer membranes of the gram-negative envelope may provide a mechanism for recruiting protein complexes to subpolar sites.

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Name: Molecular microbiology
ISSN: 1365-2958
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