Figure 2: Folding, stability and structure of design BB1.

a, In silico folding energy landscape. Each grey dot indicates the result of an independent ab initio folding calculation; black dots show results of refinement trajectories starting from design model and dark grey dots from lowest energy ab initio models. b, Size-exclusion chromatogram of the purified monomer (14 kD). c, Far-UV CD spectra at 25°C (grey line), 95°C (black dashed line) and cooled back to 25°C (black dotted line). d, Near-UV CD spectra in Tris buffer (grey line) and 7M GuHCl (black line). e, Cooperative unfolding in GuHCl monitored by near-UV CD signal at 285 nm (grey line) and tryptophan fluorescence (black line). f-j, Superpositions of the crystal structure (grey) and the design model (pink): overall backbone superposition(f); section along the β-barrel axis showing the rotameric states of core residues(g); one of the top loop with a G1 β-bulge(h); and equatorial cross-section of the β-barrel, showing the geometry of the interior volume (i) . The glycine kinks are shown as sticks. The bottom of the panel shows the cross-section of the three closest native β-barrel structures based on TM-score³³ (PDB IDs: 1JMX (0.77); 4IL6(chain O) (0.73); 1PBY (0.71)). (j) One of the bottom loops with a classic β-bulge. (k) Crystal structure and 2mFo − DFc electron density of the tryptophan corner, contoured at 1.5σ.