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Figure 1.
Fig. 1. Difference Fourier map HbI* (photoproduct) minus
HbI-CO at time delays of 5 ns and 60 µs is shown for the
entire dimer (A); CD, E, F, and heme regions of subunit A (B);
and the Phe F4 of subunit A (C). Fig. 5, which is published as
supporting information on the PNAS web site, provides equivalent
views for subunit B [the figure was produced with PYMOL (38)].
(A) A ribbon diagram of the HbI-CO dimer (gray) with side chains
for His F8 (cyan), Phe F4 (yellow), and key interface water
molecules (small cyan spheres) are shown along with the
difference Fourier map. The maps are contoured at ±3.5
 (blue and red,
respectively) for both A and B. Note the concentration of
difference density mainly in the immediate heme region and along
the F helix at 5 ns. The density distributes toward the
interface by 60 µs. Arrows (in cyan) point out the
position of two key R-state water molecules in the 5-ns map that
show clear negative density as they rapidly respond to the loss
of ligand. Removal of these two water molecules is required for
the subsequent movement of the heme groups toward the subunit
interface. (B) An  -carbon trace (gray)
for the CD region and E and F helices along with the heme group
(salmon), side chains for CD1, CD3, E7, F7, and F8, (cyan) and
F4 (yellow) are shown. The photolysis signal at the bound CO
position (labeled CO) is highly significant at 5 ns: -14 and -17
for
the A and B subunits, respectively. The strong positive feature
indicating the iron displacement (labeled Fe) is at +12 and +14
,
for the A and B subunits, respectively. Note the extensive
structural rearrangement involving the heme group at 5 ns, along
with that of the CD region and F helix. (C) Difference electron
density is shown for the region around F4 Phe at ±2.5
in
blue and red, respectively, along with the atomic model for the
liganded (salmon) and unliganded (cyan) structures. Phe F4
undergoes the largest ligand-linked side-chain rearrangement
during the R-to-T transition. As the density maps show, this
movement has not occurred at 5 ns but is completed by 60
µs after the ligand release.
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