RT Journal Article
SR Electronic
T1 Mapping the Conformational Landscape of a Dynamic Enzyme by Multitemperature and XFEL Crystallography
JF bioRxiv
FD Cold Spring Harbor Laboratory
SP 016733
DO 10.1101/016733
A1 Daniel A. Keedy
A1 Lillian R. Kenner
A1 Matthew Warkentin
A1 Rahel A. Woldeyes
A1 Jesse B. Hopkins
A1 Michael C. Thompson
A1 Aaron S. Brewster
A1 Andrew H. Van Benschoten
A1 Elizabeth L. Baxter
A1 Monarin Uervirojnangkoorn
A1 Scott E. McPhillps
A1 Jinhu Song
A1 Roberto Alonso-Mori
A1 James M. Holton
A1 William I. Weis
A1 Axel T. Brunger
A1 S. Michael Soltis
A1 Henrik Lemke
A1 Ana Gonzalez
A1 Nicholas K. Sauter
A1 Aina E. Cohen
A1 Henry van den Bedem
A1 Robert E. Thorne
A1 James S. Fraser
YR 2015
UL http://biorxiv.org/content/early/2015/08/28/016733.abstract
AB Determining the interconverting conformations of dynamic proteins in atomic detail is a major challenge for structural biology. Conformational heterogeneity in the active site of the dynamic enzyme cyclophilin A (CypA) has been previously linked to its catalytic function, but the extent to which the different conformations of these residues are correlated is unclear. Here we compare the conformational ensembles of CypA by multitemperature synchrotron crystallography and fixed-target X-ray free electron laser (XFEL) crystallography. The “diffraction-before-destruction” nature of XFEL experiments provides a radiation-damage-free view of the functionally important alternative conformations of CypA, confirming earlier synchrotron-based results. We monitored the temperature dependences of these alternative conformations with eight synchrotron datasets spanning 100-310 K. Multiconformer models show that many alternative conformations in CypA are populated only at 240 K and above, yet others remain populated or become populated at 180 K and below. These results point to a complex evolution of conformational heterogeneity between 180-240 K that involves both thermal deactivation and solvent-driven arrest of protein motions in the crystal. The lack of a single shared conformational response to temperature within the dynamic active-site network provides evidence for a conformation shuffling model, in which exchange between rotamer states for a large aromatic ring in the middle of the network shifts the conformational ensemble for the other residues in the network. Together, our multitemperature analyses and XFEL data motivate a new generation of temperature- and time-resolved experiments to structurally characterize the dynamic underpinnings of protein function.