Electrospray Ionization in Protein Studies

Many studies of biomolecules carried out by ESI-MS assume that, in the gas-phase conditions, ions retain elements of liquid-phase structures [22]. This assumption is supported by experimental results and molecular dynamics simulations. It has been suggested that solution-phase conformers of some proteins may be preserved for several tens of milliseconds after the ESI [21, 23, 24]. Because of the soft character ofESI, some protein-ligand complexes and large protein assemblies may also maintain non-covalent bonding in the gas phase [16, 24-29]. For example, nanospray electrospray ionization (nanoESI)-MS enabled the formation of complexes between a heat shock protein and an unfolding luciferase (client) to be monitored in the course of a few minutes [30]. 

A similar approach was applied in the study of holo-myoglobin [41], In both cases, the lifetimes are in good agreement with those obtained by alternative methods. Acid-induced denaturation of holo-myoglobin on a pH jump (6.5 to 3.2) coupled with ESI-MS led to observations of various protein conformations [41], The MS signals were monitored on a timescale of a few seconds with sub-second to a few seconds intervals. Some of the charge states represent lifetimes shorter than 1 s. The lifetimes obtained by TRMS and stopped-flow measurements of the Soret absorption band are convergent [41], This work pointed to 

Ionization techniques derived from ESI, such as electrosonic spray ionization (ESSI), are also suitable for studies of fragile molecules. The ESSI-MS approach enables observation of non-covalent complexes of myoglobin, protein kinase A/ATP complex, and other proteins [47]. It can be used to study on-line deprotonation reactions on peptides and proteins [48,49]. Such analyses can be done by introducing volatile bases between the ion source and mass analyzer. This method is fast one reference base can be scanned in a time interval of 1 min. The desorption electrospray ionization (DESI) technique was also implemented in the study of protein conformation in solution [50]. The interaction time between the spray solvent and the protein was estimated to be 1 ms, and it was suggested that this timescale would be too short for the studied proteins to unfold [50]. 

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