Supporting Data for "Mapping the lipid binding regions of the antifungal protein NFAP2 exploiting model membranes"

Molecular dynamics input files for the paper.

Abstract of the paper:

Fungal infections with high mortality rates represent an increasing health risk. The Neosartorya (Aspergillus) fischeri antifungal protein 2 (NFAP2) is a small, cysteine-rich, cationic protein exhibiting potent anti-Candida activity. As the underlying mechanism, pore formation has been demonstrated, however, molecular level details on its membrane disruption action are lacking. Herein we addressed lipid binding of NFAP2 using a combined computational and experimental approach on simple lipid compositions with various surface charge properties. Simulation results revealed binding preferences for negatively charged model membranes where selectivity is mediated by anionic lipid components enriched at the protein binding site but also assisted by zwitterionic lipid species. Several potential binding routes initiated by various anchoring contacts were observed, which resulted in alternative binding modes with NFAP2 residing on the membrane surface. Region 10NCPNNCKHKKG20 of the flexible N-terminal part of the protein showed potency to insert into the lipid bilayer, where the disulfide bond stabilized short motif 11CPNNC15 could play a key role, assisted by the electrostatic effects of the nearby lysines. Combined data demonstrated that the solution conformation was not perturbed markedly upon membrane interaction and the folded part of the protein also contributes to stabilizing the bound state. Data also highlighted that binding of NFAP2 to lipid vesicles is sensitively affected by environmental factors like ionic strength. Electrostatic interactions driven by anionic lipid clustering were found pivotal explaining reduced membrane activity observed at high salt conditions. Experimental data supported the lipid selective binding mechanisms, and pointed to salt-dependent effects, particularly to protein-assisted vesicle aggregation at low ionic strength. Our findings can contribute to the development of NFAP2-based anti-Candida agents and studies aiming at future medical use of peptide-based natural antifungal compounds.