Computational and cellular exploration of the protein-protein interaction between Vibrio fischeri STAS domain protein SypA and serine kinase SypE

ABSTRACT

Anti-sigma factor antagonists SpoIIAA and RsbV from Bacillus subtilis are the archetypes for single-domain STAS proteins in bacteria. The structures and mechanisms of these proteins along with their cognate anti-sigma factors have been well studied. SpoIIAA and RsbV utilize a partner-switching mechanism to regulate gene expression through protein-protein interactions to control the activity of their downstream anti-sigma factor partners. The Vibrio fischeri STAS domain protein SypA is also proposed to employ a partner-switching mechanism with its partner SypE, a serine kinase/phosphatase that controls SypA’s phosphorylation state. However, this regulation appears opposite to the canonical pathway, with SypA being the more downstream component rather than SypE. Here we explore the commonalities and differences between SypA and the canonical single-domain STAS proteins SpoIIAA and RsbV. We use a combination of AlphaFold 2 structure predictions and computational modeling to investigate the SypA-SypE binding interface. We then test a subset of our predictions in V.fischeri by generating and expressing SypA variants. Our findings suggest that, while SypA shares many sequence and structural traits with anti-sigma factor antagonist STAS domain proteins, there are significant differences that may account for SypA’s distinct regulatory output.

KEYWORDS:

• Anti-sigma factor antagonists
• biofilm
• phosphorylation
• protein-protein interaction
• STAS
• Vibrio

Acknowledgments

The authors would like to thank Xijin Lin and Prerana Shrestha for their help with biofilm experiments and strain construction, Dr. Brandon Garcia for critical review of the manuscript, and AJ Milton for editing.

Disclosure statement

No potential conflict of interest was reported by the authors.

Data availability statement

Data can be found at https://osf.io/euw4c/.

Additional information

Funding

The work was supported by the National Institutes of Health [R35 GM130355].