Crystal structures of the X-domains of a Group-1 and a Group-3 ...

J_ID: PRO Customer A_ID: PRO15 Date: 20-November-08 Stage: Page: 4
Table I. Crystallization, Data Collection, and Refinement
HCoV-229E X-domain IBV X-domain IBV X-domain
Crystallization conditions 20% PEG 8000 30% PEG 4000 1.8M (NH 4 ) 2 SO 4
0.1M Tris 0.1M Tris 0.1M Na-citrate pH 5.6
pH 8.5 pH 8.5 0.2M K-,Na-tartrate
5% MPD 0.2M MgCl 2
Data collection
Wavelength (Å) 1.04 0.808 0.808
Resolution (Å) 24.22–1.78 (1.88–1.78) 40.00–1.60 (1.64–1.60) 31.28–2.10 (2.21–2.10)
Space group P2 1 C222 1 P3 2 2
Unit-cell parameters
a (Å) 33.56 42.21 78.20
b (Å) 65.89 79.81 78.20
c (Å) 38.02 99.74 81.70
a ( ) 90 90 90
b ( ) 110.1 90 90
c ( ) 90 90 120
Solvent content (%, v/v) 33.73 43.12 66.88
Overall reflections 101,730 158,270 503,497
Unique reflections 14,479 (1946) 22,201 (1448) 17,123 (2231)
Multiplicity 4.2 (3.8) 7.1 (6.2) 11.5 (10.4)
Completeness (%) 96.9 (89.9) 97.9 (97.1) 98.4 (89.4)
R a merge (%) 5.2 (15.5) 10.3 (24.4) 7.9 (40.1)
I/r(I) 19.6 (7.8) 21.7 (6.8) 25.6 (6.5)
Refinement
Resolution (Å) 24.22–1.78 40.00–1.60 31.28–2.10
b
R cryst 0.165 0.169 0.199
b
R free 0.205 0.203 0.231
r.m.s.d. from ideal geometry
Bonds (Å) 0.013 0.011 0.022
Angles ( ) 1.284 1.349 1.896
Protein atoms 1283 1325 1335
Solvent atoms 146 212 75
Heteroatoms 20
Ramachandran plot
Most favored (%) 91.7 92.5 92.1
Additionally allowed (%) 6.9 6.8 6.6
Generously allowed (%) 0.7 0.0 0.7
Disallowed regions (%) 0.7 0.7 0.7
Values in parentheses are for the highest resolution shell.
a R merge ¼ P P
hkl i jIðhklÞ i
hIðhklÞij= P P
hkl i IðhklÞ i
, where I(hkl) is the intensity of reflection hkl and hI(hkl)i is the average
intensity over all equivalent reflections.
b R cryst ¼ P hkl jF oðhklÞ F c ðhklÞj= P hkl F oðhklÞ. R free was calculated for a test set of reflections (6%, 5%, and 6%, respectively)
omitted from the refinement.
F2
near the carboxy-terminal Lys 172 and the Phe 7-Tyr
12 segment, as well as Arg 144 and Phe 167. It could
represent two sets of twofold disordered glycerol molecules,
as glycerol was used as a cryoprotectant, but
this model was not stable during refinement. The second
unexplained density is surrounded by Leu 159-Glu
162, as well as Pro 81-Gln 88 of a symmetry-related
molecule. The third noninterpreted density is located
in the interior of the domain, close to His 45. It could
represent a second conformation of this histidine, but
this model could not be refined either.
Overall structures
The overall structures of 229E-X and IBV-X are similar
to one another, even though the sequence identity
between the two is only 22% (see Fig. 2). They are
also similar to the known structure of SARS-X (34%
and 19%, respectively). The structures show the socalled
macrodomain fold, consisting of a central,
mostly parallel b-sheet flanked by three a-helices on
one side and three on the other (see Fig. 3). Along the
polypeptide chain, the order of regular secondary
structure elements is b1-b2-a1-b3-a2-a3-b4-b5-a4-b6-
a5-b7-a6. The b-sheet topology is b1-b2-b7-b6-b3-b5-
b4, with the external strands b1 and b4 being antiparallel
to their neighbors. Interestingly, b1 is missing in
IBV-X. As a result, the positions of the N-terminal residues
are very different (15.0 Å between Leu 3 of
229E-X and the corresponding Ala 2 of IBV-X5.6) in
the two structures.
The overall r.m.s. deviation between the X-
domains of HCoV 229E and IBV (strain Beaudette) is
1.81 Å and 1.72 Å for IBV-X5.6 and IBV-X8.5, respectively,
for 146 Ca pairs out of 165 compared. The lowpH
and high-pH forms of the IBV X-domain display a
low overall r.m.s.d. of 0.46 Å, the only significant
F3
Structures of Coronaviral X-domains PROTEINSCIENCE.ORG 4
ID: srinivasanv I Black Lining: [ON] I Time: 21:11 I Path: N:/Wiley/3b2/PRO#/Vol00000/080020/APPFile/C2PRO#080020