TY - JOUR
T1 - Structural Dynamics in Ras and Related Proteins upon Nucleotide Switching
AU - Harrison, Rane A.
AU - Lu, Jia
AU - Carrasco, Martin
AU - Hunter, John
AU - Manandhar, Anuj
AU - Gondi, Sudershan
AU - Westover, Kenneth D.
AU - Engen, John R.
N1 - Funding Information:
We would like to thank Nathanael Gray for support and helpful guidance with this project, and Thomas Wales for technical assistance with HDX MS. Funding was provided by the Cancer Prevention Research Institute of Texas R1207 (KDW), ACS IRG-02-196-07 (K.D.W.), Welch Foundation I-1829 (K.D.W.), NIH R01-GM101135 (J.R.E.), a research collaboration with the Waters Corporation (J.R.E.), and the Dana Farber Cancer Institute/Northeastern University Joint Program in Cancer Drug Development (J.R.E.).
Publisher Copyright:
© 2016 Elsevier Ltd
PY - 2016/11/20
Y1 - 2016/11/20
N2 - Structural dynamics of Ras proteins contributes to their activity in signal transduction cascades. Directly targeting Ras proteins with small molecules may rely on the movement of a conserved structural motif, switch II. To understand Ras signaling and advance Ras-targeting strategies, experimental methods to measure Ras dynamics are required. Here, we demonstrate the utility of hydrogen–deuterium exchange (HDX) mass spectrometry (MS) to measure Ras dynamics by studying representatives from two branches of the Ras superfamily, Ras and Rho. A comparison of differential deuterium exchange between active (GMPPNP-bound) and inactive (GDP-bound) proteins revealed differences between the families, with the most notable differences occurring in the phosphate-binding loop and switch II. The P-loop exchange signature correlated with switch II dynamics observed in molecular dynamics simulations focused on measuring main-chain movement. HDX provides a means of evaluating Ras protein dynamics, which may be useful for understanding the mechanisms of Ras signaling, including activated signaling of pathologic mutants, and for targeting strategies that rely on protein dynamics.
AB - Structural dynamics of Ras proteins contributes to their activity in signal transduction cascades. Directly targeting Ras proteins with small molecules may rely on the movement of a conserved structural motif, switch II. To understand Ras signaling and advance Ras-targeting strategies, experimental methods to measure Ras dynamics are required. Here, we demonstrate the utility of hydrogen–deuterium exchange (HDX) mass spectrometry (MS) to measure Ras dynamics by studying representatives from two branches of the Ras superfamily, Ras and Rho. A comparison of differential deuterium exchange between active (GMPPNP-bound) and inactive (GDP-bound) proteins revealed differences between the families, with the most notable differences occurring in the phosphate-binding loop and switch II. The P-loop exchange signature correlated with switch II dynamics observed in molecular dynamics simulations focused on measuring main-chain movement. HDX provides a means of evaluating Ras protein dynamics, which may be useful for understanding the mechanisms of Ras signaling, including activated signaling of pathologic mutants, and for targeting strategies that rely on protein dynamics.
KW - Rho
KW - guanosine diphosphate (GDP)
KW - hydrogen–deuterium exchange mass spectrometry
KW - protein dynamics
KW - signal transduction
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U2 - 10.1016/j.jmb.2016.10.017
DO - 10.1016/j.jmb.2016.10.017
M3 - Article
C2 - 27751724
AN - SCOPUS:84995686979
SN - 0022-2836
VL - 428
SP - 4723
EP - 4735
JO - Journal of Molecular Biology
JF - Journal of Molecular Biology
IS - 23
ER -