Fig. 3. Presence of the open cryptic site conformation in unligated Bcl-xL.

(A) Temperature-dependent population changes in the Ile χ2, Leu χ2, Met χ3, and Val χ1 rotameric states between 288 and 308 K in apo Bcl-xL. The populations in the rotameric states were calculated from the equations shown in Materials and Methods. (B) Mapping of residues that showed substantial temperature dependence of their rotameric states. The methyl moieties with >2 SDs of rotameric population shift in response to the temperature variation are highlighted in red. (C) Van’t Hoff plot of three resonances that showed the largest temperature-dependent ¹³C chemical shift changes. (D) Methyl CPMG RD experiments of unligated Bcl-xL. The residues that showed substantial R_ex (>5 Hz) contributions to the transverse relaxation of the ¹³C methyl moiety are mapped on the structure of Bcl-xL. For those residues in the cryptic site (Leu¹⁰⁸, Ile¹¹⁴, and Leu¹⁵⁰), the RD plot and their global fitting are shown.