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fitted in the same three cavities in the N-trimer (Fig. 3). However,
when a non-inverted electron density map of N-trimer was used, both
RNA types preferred an area near the N-terminal domains of three
monomers (Fig. 4). Such a location might be of special importance as
it has been suggested that the N-terminus carries an RNA chaperone
domain.
15
The docking of the 12 nt long ssRNA or 20 nt long dsRNA
into the trimer plainly shows that the RNA spans more than one N-
monomer. Thus the RNA chaperone activity of the N protein would
require the trimer structure since single monomers apparently cannot
wrap around the RNA-panhandle or the panhandle-forming terminal
RNA sequences due to size restrictions.
We hypothesize that the N-trimer mediates panhandle dissocia-
tion in the following way:
1.
The process is started by the trimerized coil-coiled helices of the
N-terminal domain of the N protein, which recognize the dsRNA
panhandle and attach to it.
2.
The N-trimer starts to move along the panhandle, thus the end
of the panhandle goes into a gap between the middle domains of
dots per inch (Nikon LS-8000 ED). The random extensive sampling
method for indistinctive protein particles from the electron micrographs
(for the reference, see Kaukinen
et al.
, 2004
19
) was used for picking pro-
jections (~38,000) of the trimers. The top view (a) exposes the twisted struc-
ture presumably formed by the N-terminal coiled-coil helices of the
monomers.
21
The bottom view (b) exposes a ring formation in the C-termi-
nal region of the trimer presumably formed by interacting alpha-helices,
19
in
analogy with the nucleocapsid protein of rabies virus (cf. Fig. 4 in Schoehn
et al.
, 2001
29
). The two images in (c) show a stereo pair of tilted side views,
showing part of three symmetrically located connected cavities. The 3D
reconstruction was done with EMAN (http://ncmi.bcm.tmc.edu/%
7Estevel/EMAN/doc/index.html) and visualized with CHIMERA (http://
www.cgl.ucsf.edu/chimera/). Note the twisted appearance of the N-monomers
and the relatively thin junction between their N- and C-terminal domains
(arrows), which parallel the X-ray crystallography data on the N proteins of
rabies and vesicular stomatitis viruses (cf. Figs. 1 and 3 in Albertini
et al.
,
2006
13
and Green
et al.
, 2006,
14
respectively).
