Structural insights into the secretin translocation channel in the type II secretion system

Abstract

The secretin GspD of the type II secretion system (T2SS) forms a channel across the outer membrane in Gram-negative bacteria to transport substrates from the periplasm to the extracellular milieu. The lack of an atomic-resolution structure of the GspD channel hinders the investigation of substrate translocation mechanism of T2SS. Here we report cryo-EM structures of two GspD channels (∼1 MDa), from Escherichia coli K12 and Vibrio cholerae, at ∼3 Å resolution. The structures reveal a pentadecameric channel architecture, wherein three rings of GspD N domains form the periplasmic channel. The secretin domain constitutes a novel double β-barrel channel, with at least 60 β-strands in each barrel, and is stabilized by S domains. The outer membrane channel is sealed by β-strand-enriched gates. On the basis of the partially open state captured, we proposed a detailed gate-opening mechanism. Our structures provide a structural basis for understanding the secretin superfamily and the mechanism of substrate translocation in T2SS.

Multiple Structures Disclose the Secretins' Secrets

“…Based on sequence analysis, the authors previously proposed that the EPEC secretin belongs to the Vibrio-type subfamily (14). In agreement with sequence similarities, a similar structural architecture can be seen between the two Vibrio-type secretins, i.e., the EPEC structure from the work of Hay et al (6) and the Vibrio structure from the work of Yan et al (15), with loops extending from the extracellular side to form the typical cap gate (Fig. 1).…”

“…The cap gate of the Vibrio-type channel has a cone-like structure formed by 15 pairs of antiparallel ␤12-␤13 strands. The linker loops between outer ␤-barrel and cap gate (␤12 and ␤13 strands) are relatively loose, which suggests that the cap gate of the Vibrio type is probably independent of other structural parts, therefore constituting a signature distinguishing two different classes of secretins (15). The internal diameters of the cap and central gates of the secretins are not compatible with the diameters of the secreted substrates and therefore suggest that purified full-length secretins are in a "closed state" and must undergo conformational changes during the secretion process.…”

“…The internal diameters of the cap and central gates of the secretins are not compatible with the diameters of the secreted substrates and therefore suggest that purified full-length secretins are in a "closed state" and must undergo conformational changes during the secretion process. Yan et al (15) proposed that the flexible linker loops on the bottom of the cap gate may act as pivots to allow ␤-strands in the cap gate to rotate outward during substrate secretion. Regarding the central gate, formed by a four-␤-strand extension from the inner ␤-barrel, site-directed mutagenesis of the highly conserved glycine residues located at the bend points in those ␤-strands revealed their involvement as pivots to allow the conformational changes of ␤-strands in the central gate.…”

Structure and Membrane Topography of the Vibrio-Type Secretin Complex from the Type 2 Secretion System of Enteropathogenic Escherichia coli

“…S2). Analysis of other T2SS secretin structures (26) revealed that these inter-and intrasubunit salt bridges that tie the subunits together are conserved, thereby providing an explanation for the welldocumented thermal and chemical quaternary stability of secretin complexes (9).…”

“…Recently, Yan et al solved the structure of the secretin from V. cholerae (26), providing a basis to assess the similarities of the two secretins.…”