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In silico insights into the membrane binding mechanism and role of disulfide bridges in the Solanum tuberosum plant specific insert

University of British Columbia

The Solanum tuberosum plant specific insert (StPSI) has a defensive role in potato plants. The StPSI is expressed alongside an aspartic protease, which is targeted to the acidic central vacuole. It functions at an acidic pH and interacts with the anionic lipids of a membrane. The StPSI contains a set of three highly conserved disulfide bonds that bridge the protein’s helical domains, and their removal leads to enhanced membrane interactions. The present work examined several aspects of the StPSI in dimeric form, including the structure and membrane binding as a function of pH, and the effect of sequential removal of the disulfide bonds and their role in maintaining overall protein tertiary structure. Several glutamic and aspartic acid residues displayed highly perturbed pKa values, however, pH was found to have little, if any, effects on the secondary structure of the StPSI at pH 3.0 or 7.4. Coarse-grained modelling of the StPSI demonstrated poor affinity for neutral or anionic membranes at pH 7.4. Conversely, at pH 3.0, two binding modes were uncovered: Mode 1 (inactive binding) and Mode 2 (active binding). In Mode 2, N- and C-terminal residues of one monomer and numerous polar and basic residues on the second monomer interacted strongly with anionic membranes, accompanied by a re-orientation of the dimer to a more vertical position. These results offer the first examination, at near-atomic resolution, of residues mediating the StPSI-membrane interactions, and allow for the proposal of a possible fusion mechanism. Removal of disulfide bonds did not lead to destabilization of the tertiary structure at either pH 3.0 or 7.4. The StPSI possesses an extensive network of inter-monomer hydrophobic interactions and intra-monomer hydrogen bonds, which stabilize the local secondary structure, leading to a robust association between monomers, regardless of the disulfide bond state. Removal of disulfide bonds did not substantially impact secondary structure, however, they may play an important role in maintaining a less plastic structure within plant cells in order to regulate membrane affinity or targeting.

Text

2020-08-27

Attribution-NonCommercial-NoDerivatives 4.0 International

http://hdl.handle.net/2429/75716

Food Science

University of British Columbia

UBCV

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