Synchrotron X-ray footprinting as a method to visualize water in proteins

Gupta, S.; Feng, J.; Chan, L.J.G.; Petzold, C.J.; Ralston, C.Y.

doi:10.1107/S1600577516009024

Figure 8
XF-MS probes proton-coupled Zn²⁺ transfer mechanism in Zn transporter YiiP. (a) Bar plot showing radiometric water accessibility changes in response to Zn²⁺ binding measured by ratio (R) of $[^\bullet]$ OH labeling rates for residues with increase (blue), decrease (red) and no change (grey) in accessibility after rapid Zn²⁺ exposure. (b) Time courses of water accessibility changes for the indicated residues. Solid lines represent single exponential fits, which provide rate constants of conformational changes associated with Zn²⁺ binding and translocation. (c) X-ray footprinting reveals a hydrophobic gate at residue L152, which controls the opening of the inner cavity water pathway for zinc-proton exchange in the YiiP transporter. The cross-sectional view shows the position of TM helices, which separate two cavities at the intra-cellular (IC) and extra-cellular (EC) sides. Residues are color coded as in (a). XF-MS results suggest the protein conformational change alternates the membrane-facing on–off mode of zinc coordination (in D49) and protonation–deprotonation (H153) of the transport site in a coordinated fashion. (d) The red arrow indicates the proposed water pathway, which connects EC with IC after excluding the residue L152 from the surface drawing of the TM helices. Results from Gupta et al. (2014

JOURNAL OF
SYNCHROTRON
RADIATION

ISSN: 1600-5775

Volume 23| Part 5| September 2016| Pages 1056-1069

https://doi.org/10.1107/S1600577516009024

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