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Figure 5.
FIG. 5. Normalized F[o] - F[c] disaccharide electron
density map for the thioin the active site of ERManI and
comparison of the sugar ring conformations with the enzyme-bound
conformation of dMNJ in the -1 subsite and the M7 mannose
residue in the +1 subsite. A, a stereographic representation of
the difference electron density for the omitted inhibitor in the
ERManI-thiodisaccharide cocomplex. The inhibitor model is shown
to aid in map interpretation. The reducing terminal Man-  -O-CH[3]
is shown at the top, labeled as the +1 subsite residue, and the
nonreducing terminal Man residue in the 3S[1] conformation is
labeled as the -1 subsite residue. Carbon and sulfur atoms in
the structures are labeled as a reference. The electron density
map was contoured at 3  for the gray mesh and
10 for the red mesh,
demonstrating the significant electron density at the glycosidic
sulfur, the O-3' and O-4' hydroxyls of the +1 residue, and the
O-2', O-3', and O-4' hydroxyls of the -1 residue. B, the protein
structure of the ER-ManI-thiodisaccharide co-complex was aligned
with the corresponding protein structures of the
ERManI·dMNJ co-complex (20) and the co-complex of yeast
ERManI containing a Man[5]GlcNAc[2] glycan in the active site
(21) using Swiss-PdbViewer (version 3.7) (55). Displayed in the
figure are the structures of the thiodisaccharide (yellow stick
figure), dMNJ (green stick figure), and the M7 residue of the
Man[5]GlcNAc[2] glycan in the +1 subsite (white stick figure;
see Ref. 19 for oligosaccharide residue nomenclature). Carbon
and sulfur atoms in the structures are labeled as a reference.
The M7 residue is in an identical conformation as the +1 residue
of the thiodisaccharide and in a similar position except for an
offset of 0.5-0.7 Å resulting from the longer C-S bond
lengths of the thiodisaccharide. The positioning of the -1
subsite residues (dMNJ versus the -1 residue of the
thiodisaccharide) were virtually identical at the C-2, C-3, and
C-4 positions. The main differences between the two structures
were found in the equivalent of the C-1, O-5, C-5, and C-6
positions.
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