Probing protein higher-order structures by native capillary electrophoresis-mass spectrometry

Capillary electrophoresis‐mass spectrometry for protein analyses under native conditions: Current progress and perspectives

“…Stand-alone native separations are limited in terms of compositional or proteoforms information. The hyphenation of native separation techniques with MS is beneficial for obtaining information on, for example, protein identity, conformation, metal factors, or stoichiometry also for protein mixtures which has been well summarized in recent reviews [26,10,27]. The use of electromigration techniques to separate analytes under native conditions is well established since the development of blue native PAGE in 1991 [28].…”

“…In the context of native separation, the BGE also needs to maintain an analyte's HOS in solution. Therefore, BGE composition and pH need to be adjusted in approximation to the preferred physiological environment of the protein of interest [27]. This can also include the addition of specific salts which regulate some protein-protein, protein-ligand, or protein-cofactor interactions [30].…”

“…In 2019, a liquid-phase analogue of drift tube ion mobility spectrometry termed MoCE was proposed to enable ion separation as well as hydrodynamic radius and effective charge measurements in a single experiment [22]. This can be achieved by applying an electric field for separation in a laminar Poiseuille flow to incorporate Tailor dispersion analysis and electrokinetic dispersion theory [22,35,27]. Being able to determine protein charge states in solution is a great advantage as accurate hydrodynamic radii calculations rely on it.…”

Mobility capillary electrophoresis–native mass spectrometry reveals the dynamic conformational equilibrium of calmodulin and its complexes

“…[40][41][42][43][44][45][46][47][48][49] More recently, MCE has been developed and coupled together with native MS to study the ellipsoid geometric structures of proteins, as well as protein structure evolutions in different solvent environments. [50][51][52][53][54] In this study, the conformational dynamics of apo-CaM and Ca 2+ -bound CaM, as well as their binding events with a peptide ligand were investigated. Both MCE and native MS experiments were carried out for CaM samples at different Ca 2+ concentrations.…”

Recent developments and applications of capillary and microchip electrophoresis in proteomics and peptidomics (mid‐2018–2022)

“…In addition to bottom-up and top-down concepts, also a middle-down approach that involves partial digestion of proteins into longer peptides compared to the bottom-up method is used [50,51]. The ability of CZE-MS to separate complex mixtures of proteins under native conditions makes it a favorable method for native proteomics [14,[52][53][54][55][56]. Multidimensional separations combining orthogonal methods either in online or off-line mode help to achieve higher proteome or peptidome coverage [57,58].…”