Structural characterization of methionine and leucine enkephalins by hydrogen/deuterium exchange and electrospray ionization tandem mass spectrometry

Abstract

Conformational changes in two endogenous opioid active pentapeptides methionine enkephalin (Met-enk) and leucine enkephalin (Leu-enk) induced by trifluoroethanol (TFE) were identified using hydrogen/deuterium exchange (HDX), coupled with electrospray ionization (ESI) mass spectrometry. The exchange features in individual amino acid residues were characterized by acquiring tandem mass spectra of the deuterated peptides. The exact identity of the labile hydrogens involved in HDX reveals that the monomer forms of both peptides adopt an unfolded conformation in aqueous solvent, but prefer the 5-->2 beta-turn secondary structure under the membrane-mimetic environment. The ESI mass spectra of Met-enk and Leu-enk also reveal that the dimer structure of these peptides coexists with the monomer conformation. The extent of the dimer structure is dependent on the peptide concentration and nature of the solvent. The non-polar solvents facilitate the dimer formation.

On-line electrogeneration of copper-peptide complexes in microspray mass spectrometry

“…In the absence of coordination sites, the metallic ions will anchor to the amide groups of the peptide backbone via nitrogen [12]. The mass spectra obtained with a Pt electrode show the presence of the mono charged peptides at m/z ϭ 556.2 Th, the fragments b 2 , b 3 , a 4 , and b 4 , and also the formation of noncovalent dimers [44,45] Cu ϩ and Cu 2ϩ anchor sites are the same in this sequence in absence of particular copper binding sites.…”

Reaction of naphthalene-2,3-dicarboxaldehyde with enkephalins for LC-fluorescence and lc-ms analysis: Conformational studies by molecular modeling and H/D Exchange mass spectrometry

“…It is well documented in the literature that the exchange rate of a labile hydrogen depends on its solvent environment°[6,°26,°32,°33]°(i.e.,°hydrogens°involved°in hydrogen bonds or protected from the solvent are less labile than hydrogens exposed to the solvent). If the labeled Leu-Enk is eluted in the deuterated eluent, it is possible to confirm that the difference between the two peaks is due to spatial conformations and to determine them, as previously realized on°the°native°Leu-Enk° [26].…”

Protein Structural Characterization by Hydrogen/Deuterium Exchange Mass Spectrometry with Top-down Electron Capture Dissociation

“…Our data showed that the deuteration level of Cterminal fraction (short z . ions) is higher than that of the Nterminal fraction (short c ions) after 24 h full H/D exchange, which is attributed that C-terminal fraction contains a highly flexible tail (residues 71-76) and N-terminal fraction contains two hydrogen-bonded β-strands (residues 1-7 and residues 10-17) and a tightly packed α-helix (residues [23][24][25][26][27][28][29][30][31][32][33][34]. The α-helix region showed the lowest deuteration level, and the turn (residues 8-11) on the surface of the protein that connects two adjacent β-strands showed very high deuteration level.…”

“…Some reports showed some retaining of spatial deuteration patterns in HDX/CID studies. [20][21][22][23][24][25] However, others showed apparent scrambling resulted from CID in HDX/MS/MS studies. [26][27][28][29] Even though it has been reported that another threshold dissociation method, infrared multiphoton dissociation (IRMPD), 30,31 causes little or no deuterium scrambling in HDX-MS, 32 the labile modifications are lost prior to backbone cleavage, thus making it difficult to identify the modification site.…”