Inter-Domain Repulsion of Dumbbell-Shaped Calmodulin during Electrospray Ionization Revealed by Molecular Dynamics Simulations.

The mechanisms whereby protein ions are released from nanodroplets at the liquid-gas interface have continued to be controversial since electrospray ionization (ESI) mass spectrometry was widely applied in biomolecular structure analysis in solution. Several viable pathways have been proposed and verified for single-domain proteins. However, the ESI mechanism of multi-domain proteins with more complicated and flexible structures remains unclear.

Metabolite profiling and identification in living cells by coupling stable isotope tracing and induced electrospray mass spectrometry.

Direct observation of metabolites in living cells by mass spectrometry offers a bright future for biological studies but also suffers a severe challenge to untargeted peak assignment to tentative metabolite candidates. In this study, we developed a method combining stable isotope tracing and induced electrospray mass spectrometry for living-cells metabolite measurement and identification. By using C-glucose and ammonium chloride-N as the sole carbon and nitrogen sources for cell culture, Escherichia coli synthesized metabolites with N and C elements.

Native Mass Spectrometry for Peptide-Metal Interaction in Picoliter Cell Lysate.

Native mass spectrometry, which takes a high concentration of ammonium acetate (NHOAc) for ionization, coupled with tedious and solvent-consuming purification, which separates proteins from complicated environments, has shown great potential for proteins and their complexes. A high level of nonvolatile salts in the endogenous intracellular environment results in serious ion suppression and has been one of the bottlenecks for native mass spectrometry, especially for protein complexes. Herein, an integrated protocol utilizing the inner surface of a micropipette for rapid purification, desorption, and ionization of peptide-metal interaction at subfemtomole level in cell lysate was demonstrated for native mass spectrometry.

Suppression of Protein Structural Perturbations in Native Electrospray Ionization during the Final Evaporation Stages Revealed by Molecular Dynamics Simulations.

Native electrospray ionization was known to preserve the protein structure in solution, which overcame the uncontrollable acidification of droplets during transfer from solution into the gas phase in conventional electrospray ionization. However, detailed experimental studies on when and how could native electrospray ionization minimize structural perturbations remain quite unclear. Herein, we conducted molecular dynamics simulations to investigate the protein structure evolution during electrospray ionization.