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Cheung, Lennie Ka Yan

University of British Columbia

The plant-specific insert (PSI) is a membrane-active domain of some plant aspartic proteases that contributes to plant defense against pathogen infection. Despite interest in developing PSIs for crop protection or therapeutic applications, their poorly-characterized structure and function relationship has impeded a greater understanding of their mode of action. Basic and tryptophan residues have been implicated in PSI function, but a detailed investigation remains to be conducted. This study used molecular dynamics to model membrane association of the Solanum tuberosum (common potato) PSI (StPSI), identify potential design strategies for developing StPSI with improved membrane-fusing efficiency, and gain insights on the inactive, closed monomer structure. Using the membrane-active dimeric StPSI structure as a template, 8 variants with substitutions to key basic or tryptophan residues were investigated. Wild-type StPSI was predicted to have a flexible hydrophobic core mediated by the tryptophan (W) at position 18 (W18), and have dynamic structural and surface properties facilitated by the formation and dissolution of inter-helical salt bridges. The bilayer association of wild-type StPSI was predicted to involve three steps: initial anchoring of specific domains, dimer reorientation and gradual descent to the bilayer surface facilitated by hydrophobic interactions and electrostatic attraction. Restricted anchoring, impaired dimer reorientation, and the lack of a specific salt bridge were attributes observed in StPSI variants with increased predicted tendency to maintain more vertical orientations on the bilayer, a characteristic hypothesized to increase fusion efficiency. The relationship between these three attributes and the predicted initial orientation of StPSI on model bilayers was further supported by computational analysis of two additional StPSI variants designed using the identified attributes as guiding principles, suggesting the potential utility of these attributes as design principles for developing StPSI with enhanced fusogenic efficiency. Domain swapping involving a hinge domain, identified as a region of interest, was proposed as a possible mechanism of StPSI dimerization. The presence of W18 may further induce asymmetry in the dimerized assembly, thereby influencing StPSI function. These findings deepened current understanding of the StPSI structure and function relationship and may facilitate development of the PSI for novel applications ranging from plant biotechnology to human health.

Text

2024-07-10

Attribution-NonCommercial-NoDerivatives 4.0 International

http://hdl.handle.net/2429/88609

Food Science

University of British Columbia

UBCV

http://creativecommons.org/licenses/by-nc-nd/4.0/