The Structure of Cyclic Neuropeptide Somatostatin and Octapeptide Octreotide in the Presence of Copper Ions: Insights from Transition Metal Ion FRET and Native Ion Mobility-Mass Spectrometry

Abstract

The conformation and function of somatostatin (SST), a cyclic neuropeptide, was recently found to be altered in the presence of Cu(II) ions, which leads to self-aggregation and loss of biological function as a neurotransmitter. However, the impact of Cu(II) ions on the structure and function of SST is not fully understood. In this work, transition metal ion Förster resonance energy transfer (tmFRET) and native ion mobility-mass spectrometry (IM-MS) were utilized to study the structures of well-defined gas-phase ions of SST and of a smaller analogue, octreotide (OCT). The tmFRET results suggest two binding sites of Cu(II) ions in both native-like SST and OCT ions, either in close proximity to the disulfide bond or complexed by two aromatic residues, consistent with results obtained from collision-induced dissociation (CID). The former binding site was reported to initiate aggregation of SST, while the latter binding site could directly affect the essential motif for receptor binding and therefore impair the biological function of SST and OCT when bound to SST receptors. Our results demonstrate that tmFRET is capable of locating transition metal ion binding sites in neuropeptides. Furthermore, multiple distance constraints (tmFRET) and global shape (IM-MS) provide additional structural insights of SST and OCT ions upon metal binding, which is related to the self-aggregation mechanisms and overall biological functions.

Transition metal ions and neurotransmitters: coordination chemistry and implications for neurodegeneration

“…It should be noted that other Cu( ii )-binding sites, in addition to the N-terminal region, have also been suggested within the sidechain of Phe6 and Phe7 via cation–π interactions. 165 …”

Probing the Stability of a β-Hairpin Scaffold after Desolvation

“…Recently, we developed an approach based on IM-MS and FRET to obtain complementary overall shape and intramolecular distance information on biomolecular ions, ,, which in combination with computational models yields the most plausible gas-phase structures until now. In IM-MS ions traversing a tube filled with neutral gas can be separated based on their charge, size and interaction with the drift gas.…”

“…Förster resonance energy transfer (FRET) is a process that occurs when the emission spectrum of an excited donor fluorophore overlaps with the absorption spectrum of an acceptor chromophore . The resulting radiationless dipole–dipole interaction between the donor and acceptor is strongly distance dependent and allows one to obtain conformational information based on strategically positioned labeling sites. , Recently, customization of mass spectrometers has enabled FRET-based studies of mass-selected species to be carried out. − Transition metal ion FRET (tmFRET), a recent expansion of FRET, employs transition metal ions, e.g. Cu­(II), as acceptor chromophores. , Due to reduced spectral overlap, the working range is significantly shorter (10–40 Å) compared to traditional FRET (40–100 Å).…”

“…Cu­(II), as acceptor chromophores. , Due to reduced spectral overlap, the working range is significantly shorter (10–40 Å) compared to traditional FRET (40–100 Å). Recent studies have applied gas-phase tmFRET to helical and cyclic peptides and were able to obtain donor–acceptor distances (r DA ) as low as 14.9 Å, ,, which promotes tmFRET as a promising avenue to probe short interstrand distances within the β-hairpin model.…”

“…Also, the dipole–dipole interaction between the fluorophore and the Cu­(II) ion may be restricted due to polarization of the π-electron cloud of the dihistidine motif upon Cu­(II) complexation . Consequently, the R 0 value may be overestimated when Cu­(II) is bound by aromatic residues, which results in larger r DA values. , …”