A dodecameric ring-like structure of the N0 domain of the type II secretin from enterotoxigenic Escherichia coli

Abstract

In many bacteria, secretins from the type II secretion system (T2SS) function as outer membrane gated channels that enable passage of folded proteins from the periplasm into the extracellular milieu. Cryo-electron microscopy of the T2SS secretin GspD revealed previously the dodecameric cylindrical architecture of secretins, and crystal structures of periplasmic secretin domains showed a modular domain organization. However, no high-resolution experimental data has as yet been provided about how the entire T2SS secretin or its domains are organized in a cylindrical fashion. Here we present a crystal structure of the N0 domain of the T2SS secretin GspD from enterotoxigenic Escherichia coli containing a helix with 12 subunits per turn. The helix has an outer diameter of ~125 Å and a pitch of only 24 Å which suggests a model of a cylindrical dodecameric N0 ring whose dimensions correspond with the cryo-electron microscopy map of Vibrio cholerae GspD. The N0 domain is known to interact with the HR domain of the inner membrane T2SS protein GspC. When the new N0 ring model is combined with the known N0• HR crystal structure, a dodecameric double-ring of twelve N0-HR heterodimers is obtained. In contrast, the previously observed compact N0-N1 GspD module is not compatible with the N0 ring. Interestingly, a N0-N1 T3SS homolog is compatible with forming a N0-N1 dodecameric ring, due to a different N0-vs-N1 orientation. This suggests that the dodecameric N0 ring is an important feature of T2SS secretins with periplasmic domains undergoing considerable motions during exoprotein translocation.

Secretion systems in Gram-negative bacteria: structural and mechanistic insights

“…24). The 19 Å cryo-EM reconstruction of the V. cholerae T2SS secretin revealed a closed cylindrical structure with an outer diameter of 155 Å and a length of 200 Å 24 , into which the crystal structure of the N0-N1-N2 domains could be fitted [24][25][26] (FIG. 1b).…”

The role of intrinsic disorder and dynamics in the assembly and function of the type II secretion system

“…An anti-parallel two αhelical core (α1-α2) is packed between an anti-parallel β1-β3 sheet on one side and an anti-parallel β2-β5-β4 sheet on the other (Fig. 2C) [71,72]. The N 1 , N 2 and N 3 domains on the other hand share structural homology with the KH-domain motif and are composed of a three stranded anti-parallel β-sheet (N 1 :β6-β8-β7; N 2 :β9-β11-β10; N 3 :β12-β14-β13) packed against two α-helices (N 1 :α3-α4; N 2 :α5-α6; N 3 :α7-α8) ( Fig.…”

The trans‐envelope architecture and function of the type 2 secretion system: new insights raising new questions

“…The N0‐, N1‐, N2‐ and N3‐domains have the most divergent sequences. The crystal and solution structures of one or more of these domains have been resolved alone (Korotkov et al ., ; Van der Meeren et al ., ) or in complex with partners (Korotkov et al ., 2009b; 2011; Gu et al ., ). None of them docked correctly to the cryo‐EM secretin structure (Yan et al ., ) suggesting differences between native domain arrangement and crystal packing.…”