Journal: Structure / Year: 2005 Title: Features of reovirus outer capsid protein mu1 revealed by electron cryomicroscopy and image reconstruction of the virion at 7.0 Angstrom resolution. Authors: Xing Zhang / Yongchang Ji / Lan Zhang / Stephen C Harrison / Dan C Marinescu / Max L Nibert / Timothy S Baker / Abstract: Reovirus is a useful model for addressing the molecular basis of membrane penetration by one of the larger nonenveloped animal viruses. We now report the structure of the reovirus virion at ...Reovirus is a useful model for addressing the molecular basis of membrane penetration by one of the larger nonenveloped animal viruses. We now report the structure of the reovirus virion at approximately 7.0 A resolution as obtained by electron cryomicroscopy and three-dimensional image reconstruction. Several features of the myristoylated outer capsid protein mu1, not seen in a previous X-ray crystal structure of the mu1-sigma3 heterohexamer, are evident in the virion. These features appear to be important for stabilizing the outer capsid, regulating the conformational changes in mu1 that accompany perforation of target membranes, and contributing directly to membrane penetration during cell entry.
A: major outer-capsid protein mu1 B: major outer-capsid protein mu1 C: major outer-capsid protein mu1 S: major capsid surface protein sigma-3 D: major capsid surface protein sigma-3 E: major capsid surface protein sigma-3 F: major capsid surface protein sigma-3 M: major capsid surface protein sigma-3 N: major capsid surface protein sigma-3 O: major capsid surface protein sigma-3 G: major capsid surface protein sigma-3 H: major capsid surface protein sigma-3 I: major capsid surface protein sigma-3 P: major outer-capsid protein mu1 Q: major outer-capsid protein mu1 R: major outer-capsid protein mu1 J: major outer-capsid protein mu1 K: major outer-capsid protein mu1 L: major outer-capsid protein mu1 T: major outer-capsid protein mu1 U: guanylyltransferase V: major core protein lambda 1 W: major core protein lambda 1 X: Sigma 2 protein Y: Sigma 2 protein Z: Sigma 2 protein 1: Minor core protein lambda 3
A: major outer-capsid protein mu1 B: major outer-capsid protein mu1 C: major outer-capsid protein mu1 S: major capsid surface protein sigma-3 D: major capsid surface protein sigma-3 E: major capsid surface protein sigma-3 F: major capsid surface protein sigma-3 M: major capsid surface protein sigma-3 N: major capsid surface protein sigma-3 O: major capsid surface protein sigma-3 G: major capsid surface protein sigma-3 H: major capsid surface protein sigma-3 I: major capsid surface protein sigma-3 P: major outer-capsid protein mu1 Q: major outer-capsid protein mu1 R: major outer-capsid protein mu1 J: major outer-capsid protein mu1 K: major outer-capsid protein mu1 L: major outer-capsid protein mu1 T: major outer-capsid protein mu1 U: guanylyltransferase V: major core protein lambda 1 W: major core protein lambda 1 X: Sigma 2 protein Y: Sigma 2 protein Z: Sigma 2 protein 1: Minor core protein lambda 3
A: major outer-capsid protein mu1 B: major outer-capsid protein mu1 C: major outer-capsid protein mu1 S: major capsid surface protein sigma-3 D: major capsid surface protein sigma-3 E: major capsid surface protein sigma-3 F: major capsid surface protein sigma-3 M: major capsid surface protein sigma-3 N: major capsid surface protein sigma-3 O: major capsid surface protein sigma-3 G: major capsid surface protein sigma-3 H: major capsid surface protein sigma-3 I: major capsid surface protein sigma-3 P: major outer-capsid protein mu1 Q: major outer-capsid protein mu1 R: major outer-capsid protein mu1 J: major outer-capsid protein mu1 K: major outer-capsid protein mu1 L: major outer-capsid protein mu1 T: major outer-capsid protein mu1 U: guanylyltransferase V: major core protein lambda 1 W: major core protein lambda 1 X: Sigma 2 protein Y: Sigma 2 protein Z: Sigma 2 protein 1: Minor core protein lambda 3
x 5
icosahedral pentamer
9.44 MDa, 135 polymers
Theoretical mass
Number of molelcules
Total (without water)
9,439,629
135
Polymers
9,439,629
135
Non-polymers
0
0
Water
0
Type
Name
Symmetry operation
Number
identity operation
1_555
x,y,z
1
point symmetry operation
4
4
A: major outer-capsid protein mu1 B: major outer-capsid protein mu1 C: major outer-capsid protein mu1 S: major capsid surface protein sigma-3 D: major capsid surface protein sigma-3 E: major capsid surface protein sigma-3 F: major capsid surface protein sigma-3 M: major capsid surface protein sigma-3 N: major capsid surface protein sigma-3 O: major capsid surface protein sigma-3 G: major capsid surface protein sigma-3 H: major capsid surface protein sigma-3 I: major capsid surface protein sigma-3 P: major outer-capsid protein mu1 Q: major outer-capsid protein mu1 R: major outer-capsid protein mu1 J: major outer-capsid protein mu1 K: major outer-capsid protein mu1 L: major outer-capsid protein mu1 T: major outer-capsid protein mu1 U: guanylyltransferase V: major core protein lambda 1 W: major core protein lambda 1 X: Sigma 2 protein Y: Sigma 2 protein Z: Sigma 2 protein 1: Minor core protein lambda 3
x 6
icosahedral 23 hexamer
11.3 MDa, 162 polymers
Theoretical mass
Number of molelcules
Total (without water)
11,327,554
162
Polymers
11,327,554
162
Non-polymers
0
0
Water
0
Type
Name
Symmetry operation
Number
identity operation
1_555
x,y,z
1
point symmetry operation
5
5
Idetical with deposited unit in distinct coordinate
icosahedral asymmetric unit, std point frame
Type
Name
Symmetry operation
Number
transform to point frame
1
Symmetry
Point symmetry: (Hermann–Mauguin notation: 532 / Schoenflies symbol: I (icosahedral))
-
Components
-
Protein , 6 types, 27 molecules ABCPQRJKLTSDEFMNOGHIUVWXYZ1
#1: Protein
majorouter-capsidproteinmu1 / Coordinate model: Cα atoms only
Mass: 76346.336 Da / Num. of mol.: 10 / Source method: isolated from a natural source / Source: (natural) Mammalian orthoreovirus 1 / Genus: Orthoreovirus / Species: Mammalian orthoreovirus / Strain: Lang / References: UniProt: P11077
#2: Protein
majorcapsidsurfaceproteinsigma-3 / Coordinate model: Cα atoms only
Mass: 41237.117 Da / Num. of mol.: 10 / Source method: isolated from a natural source / Source: (natural) Mammalian orthoreovirus 1 / Genus: Orthoreovirus / Species: Mammalian orthoreovirus / Strain: Lang / References: UniProt: P07939
#3: Protein
guanylyltransferase / Coordinate model: Cα atoms only
Mass: 144098.766 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Mammalian orthoreovirus 3 / Genus: Orthoreovirus / Species: Mammalian orthoreovirus / Strain: Dearing / References: UniProt: P11079
#4: Protein
majorcoreproteinlambda1 / Coordinate model: Cα atoms only
Mass: 142008.359 Da / Num. of mol.: 2 / Source method: isolated from a natural source / Source: (natural) Mammalian orthoreovirus 1 / Genus: Orthoreovirus / Species: Mammalian orthoreovirus / Strain: Lang / References: UniProt: Q9WAB2
#5: Protein
Sigma2protein / Core protein / Coordinate model: Cα atoms only
Mass: 47155.211 Da / Num. of mol.: 3 / Source method: isolated from a natural source / Source: (natural) Mammalian orthoreovirus 1 / Genus: Orthoreovirus / Species: Mammalian orthoreovirus / Strain: Lang / References: UniProt: P11314
#6: Protein
Minorcoreproteinlambda3 / Coordinate model: Cα atoms only
Mass: 142510.062 Da / Num. of mol.: 1 / Source method: isolated from a natural source / Source: (natural) Mammalian orthoreovirus 1 / Genus: Orthoreovirus / Species: Mammalian orthoreovirus / Strain: Lang / References: UniProt: P17376, UniProt: P0CK32*PLUS
-
Experimental details
-
Experiment
Experiment
Method: ELECTRON MICROSCOPY
EM experiment
Aggregation state: PARTICLE / 3D reconstruction method: single particle reconstruction
-
Sample preparation
Component
Name: HUMAN REOVIRUS VIRIONS (T3D) / Type: VIRUS / Details: The structure was monodisperse.
Details of virus
Host category: MAMMALIAN
Buffer solution
Name: 10 mM TRIS / pH: 7.5 / Details: 10 mM TRIS
Specimen
Conc.: 2 mg/ml / Embedding applied: NO / Shadowing applied: NO / Staining applied: NO / Vitrification applied: YES
Specimen support
Details: HOLEY CARBON
Vitrification
Cryogen name: ETHANE / Details: PLUNGED INTO ETHANE
-
Electron microscopy imaging
EM imaging
Electron source: FIELD EMISSION GUN / Illumination mode: FLOOD BEAM / Mode: BRIGHT FIELD / Specimen-ID: 1
ID
Accelerating voltage (kV)
Calibrated magnification (X)
Date
Details
Model
Cs (mm)
Nominal defocus max (nm)
Nominal defocus min (nm)
Nominal magnification (X)
Temperature (K)
Tilt angle max (°)
Tilt angle min (°)
1
200
45000
Dec 10, 1997
SAMPLES WERE MAINTAINED AT -176 CELSIUS DEGREE IN THE ELECTRON MICROSCOPES.
FEI/PHILIPS CM200FEG
2
3200
1300
38000
97
0
0
2
300
47440
Apr 18, 2002
FEI/PHILIPS CM300FEG/T
39200
Image recording
Electron dose: 20 e/Å2 / Film or detector model: KODAK SO-163 FILM
Image scans
Num. digital images: 54
-
Processing
CTF correction
Details: CTF correction of each particle
Symmetry
Point symmetry: I (icosahedral)
3D reconstruction
Method: PFT, OOR, POR / Resolution: 7 Å / Num. of particles: 7939 / Nominal pixel size: 2.3 Å / Actual pixel size: 2.21 Å / Magnification calibration: CROSS-CORRELATION / Symmetry type: POINT
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