Fig. 3: Comparison of Anc4 + Loop II and Anc5ΔLoop I exhibit the structural and dynamics contribution of Loop II and scaffold substitutions.

A The difference in Δroot mean square fluctuation (ΔRSMF = RMSF of Anc4 + Loop II – RMSF of Anc5ΔLoop I) is mapped on the structure of Anc5ΔLoop I as the thickness of the cartoon representation. Spheres indicates two catalytic glutamic acids and one serine that holds catalytic water. B Plots of the RMSF of Cα of each residue in Anc4 + Loop II (blue) and Anc5ΔLoop I (orange). Error bars are indicated as shades (n = 4). A schematic representation of secondary structures of GH19 chitinase is shown. Dashed squares indicate the region where ΔRSMF is more than 0.05 nm. RMSDs of the Cα atoms of all residues are shown in Supplementary Fig. 5B.C Cartoon_tube representation of Anc5. Sphere representation indicates 45 substitution residues (purple) and two catalytic glutamic acids and one serine that holds catalytic water (magenta). Resides around 4 Å from loop II are shown in sticks. Intramolecular interactions stabilizing loop II regions are shown in the enlargements. Resides in Anc4 and Anc5 state are shown in gray and purple sticks, respectively. Black dashed lines indicate hydrogen bonds. Residue numbers are based on Anc4. In the case that the number shifts due to loop insertions, residue numbers of Anc5 state are in parentheses. An asterisk indicates the number of Anc4 + Loop II. Loop regions I to VI are shown in red, green, light blue, purple, orange, and cyan, respectively. D Left, computational analysis of long-range communication in Anc5ΔLoop I. Cartoon representation of Anc5ΔLoop I with cartoon_tube representation of loop II to VI (shown in green, slate, purple, orange, and cyan, respectively). Sphere representation indicates 45 substitutions residues (purple) and two glutamic acids and one serine that holds catalytic water (magenta). Right, rigidity-transmission allostery (RTA) communication analysis on Anc5ΔLoop I. Residues are colored based on the intensity (red being highest) of long-range rigidity-transmission communication with loop II (green). Spheres indicate 45 substitutions.