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Figure 1.
Conformational change in actin subdomain 2 on phosphorylation
of Tyr-53. (A and B) Close views of subdomain 2 in the
structures of unphosphorylated actin and pY53-actin, showing
omit electron density maps (contoured at 1 σ) around Tyr-53
(see Figs. S3 and S5 for a full view of the G1-actin structure).
The D-loop was not visualized in the unphosphorylated structure.
Hydrogen-bonding contacts (red dashed lines) between the oxygen
atoms of the phosphate group on Tyr-53 and residues of the
D-loop stabilize the conformation of the D-loop in the structure
of pY53-actin. This and other figures of the paper were
generated with the program PyMOL
(http://pymol.sourceforge.net/). (C) Phosphorylation protects
the D-loop from subtilisin cleavage, as shown by the ≈50%
decrease in the initial rate of digestion. (D and E) Based on
the increase in fluorescence as etheno-ATP replaces actin-bound
ATP, phosphorylation reduces the rate of nucleotide exchange
from 0.011 s^−1 for unphosphorylated actin to 0.006 s^−1 for
pY53-actin, but profilin accelerates and gelsolin inhibits
nucleotide exchange to the same extents for both forms of actin.
The increase in fluorescence at equilibrium for pY53-actin is
only 50% of the increase for unphosphorylated actin. Data were
recorded every 10 s.
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