Figure 1.
Structure of synthetic open-chain MCoTI, structural overview and fragment ion formation.
(A) 3D structure of synthetic open-chain cystine knot oMCoTI (pdb: 2IT8).15 Active loop is shown in red. (B) A 6 Å close-up on the disulfide-tightened core of MCoTI. For both (A) and (B) disulfides are shown in yellow. (C) Overview of the generation of peptidic fragment ions upon CID. (D) Sequences of the parent MCoTI wild type (wt) and the miniproteins (1–3) used in this study. Positions with altered amino acid residues (regarding the wild type sequence) are marked red.
Figure 2.
(A) CID of the doubly charged ion of reduced peptide 1. (B) CID of the doubly charged ion of folded miniprotein. (C) CID of the five-fold charged ion of folded miniprotein.
Figure 3.
MS3 of the CID-obtained major fragments of MCoTI 1.
(A) MS3 of c10 of the doubly charged ions of unfolded peptide. (B) MS3 of c16y19 of the doubly-charged ions of folded miniprotein. (C) MS3 of y14 of the doubly charged ions of unfolded peptide. (D) MS3 of y24 of the five-fold charged ions of folded miniprotein. Arrows above the spectra indicate intensity amplifications.
Figure 4.
MS3 of y14-32 (A) and y14+32 (B) and the resulting combinatorial interpretation.
In red are inexistent peaks to provide evidence on the respective Ptc or Dha cleavage. Arrows above the spectra indicate intensity amplifications.
Figure 5.
Spectra and assigned structures of the bridged fragments.
(A) MS3 of the bridged fragment of 1. (B) MS3 of the bridged fragment of 2. Arrows above the spectra indicate intensity amplifications.
Table 1.
Amount of loop ratios in CID of the respective peptides.
Figure 6.
Optimization of the collision energy towards a maximum formation of fragment c10y26 of 2.