Figure 5. Dynamic folding modulation design and structural analysis of FGF21^SS variant.

1. Sequence alignment of FGF21, FGF21^SS, and FGF19 in the mutated region. The mutation sites are indicated by boxes. Cysteines for disulfide bond formation are highlighted on a yellow background. The secondary structure of FGF21 is shown on the top. The N‐terminal residue IDs of FGF21^SS are indicated under the sequence. # is used to identify the mutated residues (codes are colored blue) and the same residues having different sequence number in FGF21^SS from wild type.
2. 2D ¹H‐¹⁵N HSQC spectra comparison of FGF21^core (black) and FGF21^SS/core (red) revealing the elimination of the loop conformation in FGF21^SS. Assignments for FGF21^core and FGF21^SS/core are labeled as in Figs 2B and 5A, respectively.
3. Temperature denaturation CD experiments indicating the significant increase of thermostability. CD value was recorded and normalized at 222 nm. T_m value of the proteins is fitted using Boltzmann function.
4. Superimposition of FGF21^SS/core (skyblue) and FGF21^core (green) structure ensembles (10 energy‐refined structures each) in ribbon representation showing the structure conservation upon SS bond mutation.
5. Cartoon representation of FGF21^SS/core. Cys31^#‐Cys43^# (between β2 and β3) and Cys75‐Cys93 (between β6 and β9) SS bonds are highlighted and shown as stick representations.