Chaperone Priming of Pilus Subunits Facilitates a Topological Transition that Drives Fiber Formation

Abstract

Periplasmic chaperones direct the assembly of adhesive, multi-subunit pilus fibers that play critical roles in bacterial pathogenesis. Pilus assembly occurs via a donor strand exchange mechanism in which the N-terminal extension of one subunit replaces the chaperone G(1) strand that transiently occupies a groove in the neighboring subunit. Here, we show that the chaperone primes the subunit for assembly by holding the groove in an open, activated conformation. During donor strand exchange, the subunit undergoes a topological transition that triggers the closure of the groove and seals the N-terminal extension in place. It is this topological transition, made possible only by the priming action of the chaperone that drives subunit assembly into the fiber.

Quality control of disulfide bond formation in pilus subunits by the chaperone FimC

“…Recently, we reported the NMR structure of a designed, selfcomplemented FimA variant (FimAa) 28 , in which FimA is artificially extended at its C terminus by a hexaglycine linker followed by the FimA donor strand segment (residues [1][2][3][4][5][6][7][8][9][10][11][12][13][14][15][16][17][18][19][20]. FimAa has the same slow, spontaneous folding rate as wild-type FimA (1.6-h folding half-life) and adopts a conformation in which the C-terminal copy of the donor strand is incorporated into the tertiary structure in an antiparallel orientation relative to the FimA F strand, which corres ponds to the expected donor strand insertion pattern in the quaternary structure of the pilus rod 28 .…”

“…Each subunit has an incomplete immunoglobulin-like pilin fold 1 lacking the C-terminal β-strand. In addition, each subunit (except for FimH) has an N-terminal extension ('donor strand'), which is donated to the preceding subunit in the pilus, thereby completing its fold 1,6,7 .…”

“…1b). The chaperonesubunit complexes then diffuse to the outer membrane usher FimD, where the N-terminal extension of the incoming subunits replaces the donor strand of the chaperone on the previously incorporated subunit at the growing pilus end in a reaction termed donor strand exchange 7,11,[14][15][16] . In contrast to the 'parallel' orientation of the donor strand in FimC-subunit complexes, the donor strand complementing the fold of neighboring subunits in the pilus has an antiparallel orientation relative to the F strand of the acceptor subunit.…”

Rationally designed small compounds inhibit pilus biogenesis in uropathogenic bacteria

“…Pilus subunits consist of a single, incomplete Ig-like fold that lacks its C-terminal seventh (G) ␤-strand (12,13). In a process termed donor-strand complementation, the chaperone's G1 ␤-strand provides in trans the pilin's seventh strand (12)(13)(14)(15). The mechanism of action of PapD depends on the formation of an ion pair between the C-terminal carboxyl group of each subunit and the conserved chaperone cleft residues R8 and K112 (7,8).…”

“…Assembly occurs by a process termed donor-strand exchange, in which the G1 ␤-strand of the chaperone is replaced by an N-terminal extension that is present on every subunit (14,15,18). Upon exchange, the pilus subunit undergoes a topological transition that triggers the closure of its groove, incorporating its neighbor's N-terminal extension as part of its own Ig fold (15,19).…”