Adhesive pili of the chaperone-usher family - Escherichia coli: Pathotypes and Principles of Pathogenesis - page 360

plug switch induces a conformational change in the β-barrel domain, shifting
the shape of the channel pore from ovular (52 Å × 28 Å) to nearly circular
(44 Å × 36 Å) ( Phan et al., 2011 ). This in turn allows a range of subunit diam-
eters (∼20-25 Å) to pass through unobstructed. The plug then docks onto the
NTD, as suggested in the crystal structure of the FimD usher ( Phan et al., 2011 )
and from work in the Pap system ( Volkan et al., 2012 ). The plug-NTD inter-
action results in the usher adopting an open state.
NTD and CTDs, which reside in the periplasmic space, serve as the work-
horses of assembly within this molecular machine. Functional studies directly
implicate the periplasmic domains of the usher in catalysis of pilus forma-
tion, as mutations in either NTD or CTD ( Henderson et al., 2011 ; Thanassi
et al., 2002 ) and deletions of the plug ( Huang et al., 2009 ; Mapingire et al.,
2009 ) abrogate assembly. To better dissect the role of each domain, affini-
ties were measured for the interactions of chaperone-subunit complexes with
purified NTD, CTD2, and plug domains from the PapC usher. NTD only
binds the chaperone-adhesin complex with high affinity (K D = 1 nM) and the
chaperone-rod adaptor complex with much weaker affinity (K D = 1 µM); plug
and the plug-NTD complex bind the chaperone and all chaperone-subunit
complexes with equal affinity (K D = 10-100 nM); CTD2 binds chaperone
and all chaperone-subunit complexes with equal affinity (K D = 1 µM), but
lacks affinity for the chaperone-terminator complex; and CTD2 dislocates
the PapD-PapG complex from the NTD of the PapC usher ( Volkan et al.,
2012 ). These data suggest that (a) the tight-binding NTD serves as the initial
anchoring site for the chaperone-adhesin complex, (b) plug or the NTD-
plug complex serves as the docking site for all other chaperone-subunit com-
plexes, and (c) CTD2 likely dissociates chaperone-subunits from NTD and
plug, with the exception of the chaperone-terminator complex, which docks
on plug or the NTD-plug complex and halts further pilus growth.
It is interesting to note that NTD can discriminate between subunits loaded
onto the chaperone, despite the high structural homology that pilins share.
Analogously to PapC NTD, the FimD NTD is selective; isothermal titration
calorimetry (ITC) experiments show that FimD NTD binds the chaperone when
the chaperone is loaded with FimH or FimF but not with FimG or FimA ( Nishi-
yama et al., 2003 ). The molecular basis for this selectivity may be indirectly
inferred from the crystal structures of the FimD NTD-FimC-FimH complex
( Nishiyama et al., 2005 ) and FimD NTD-FimC-FimF complex ( Eidam et al.,
2008 ). In these crystal structures, FimD NTD consists of two flexible N- and
C-terminal tails and a uniquely folded core, in which a strained, three-stranded,
antiparallel β-sheet and a two-stranded, antiparallel β-sheet pack against one
another and are bridged by a segment containing a 3 10 -helix and two α-helices.
These structures along with supporting NMR data reveal that the intrinsically
disordered 24-residue N-terminal tail of NTD adopts an ordered conforma-
tion upon binding. As expected, deletion of the N-terminal tail of NTD com-
pletely abolishes pilus assembly in vivo ( Nishiyama et al., 2005 ). In the ternary
complex, the N-terminal tail of NTD contacts the chaperone and the subunit