Early Structural Evolution of Native Cytochrome c after Solvent Removal

Graphical Abstract

Electrostatic stabilization: When a native protein structure is exposed to vacuum as in native electrospray ionization (ESI), what are the very early structural changes caused by desolvation? Molecular dynamics (MD) calculations show that before global structural changes occur, the native fold can be transiently stabilized by the formation of salt bridges and ionic hydrogen bonds on the protein surface (see scheme).

Abstract

Electrospray ionization transfers thermally labile biomolecules, such as proteins, from solution into the gas phase, where they can be studied by mass spectrometry. Covalent bonds are generally preserved during and after the phase transition, but it is less clear to what extent noncovalent interactions are affected by the new gaseous environment. Here, we present atomic-level computational data on the structural rearrangement of native cytochrome c immediately after solvent removal. The first structural changes after desolvation occur surprisingly early, on a timescale of picoseconds. For the time segment of up to 4.2 ns investigated here, we observed no significant breaking of native noncovalent bonds; instead, we found formation of new noncovalent bonds. This generally involves charged residues on the protein surface, resulting in transiently stabilized intermediate structures with a global fold that is essentially the same as that in solution. Comparison with data from native electron capture dissociation experiments corroborates both its mechanistic postulations and our computational predictions, and suggests that global structural changes take place on a millisecond timescale not covered by our simulations.