Structural and Functional Studies on the Interaction of GspC and GspD in the Type II Secretion System

Abstract

Type II secretion systems (T2SSs) are critical for secretion of many proteins from Gram-negative bacteria. In the T2SS, the outer membrane secretin GspD forms a multimeric pore for translocation of secreted proteins. GspD and the inner membrane protein GspC interact with each other via periplasmic domains. Three different crystal structures of the homology region domain of GspC (GspCHR) in complex with either two or three domains of the N-terminal region of GspD from enterotoxigenic Escherichia coli show that GspCHR adopts an all-β topology. N-terminal β-strands of GspC and the N0 domain of GspD are major components of the interface between these inner and outer membrane proteins from the T2SS. The biological relevance of the observed GspC–GspD interface is shown by analysis of variant proteins in two-hybrid studies and by the effect of mutations in homologous genes on extracellular secretion and subcellular distribution of GspC in Vibrio cholerae. Substitutions of interface residues of GspD have a dramatic effect on the focal distribution of GspC in V. cholerae. These studies indicate that the GspCHR–GspDN0 interactions observed in the crystal structure are essential for T2SS function. Possible implications of our structures for the stoichiometry of the T2SS and exoprotein secretion are discussed.

Structure and assembly of an inner membrane platform for initiation of type IV pilus biogenesis

“…It is reasonable to infer that the formation of secretin dodecamers in the outer membrane could act as a driver for oligomerization of the PilMNO inner membrane components. Indeed, similar suggestions have been made in the case of the T2SS (34). The consequence would be that, when a major pilin binds to this assembly, pilins from adjacent PilMNO complexes are brought into close spatial proximity, where they could associate and thus be immediately available for incorporation into a pilus fiber.…”

“…The T2SS secretin GspD and the T4P secretin PilQ form dodecameric assemblies enclosing chambers through which the secreted proteins pass (8, 32). Recently, it has been noted that the lipoprotein PilP, which is located in the inner membrane and binds to PilQ (33), adopts a similar β structure to GspC, a component of the T2SS that has a related function in binding to its cognate secretin GspD (34). Moreover, there are also similarities in the ways the two secretin-binding proteins recognize the secretin α/β domains in PilQ and GspD (8,34).…”

“…Recently, it has been noted that the lipoprotein PilP, which is located in the inner membrane and binds to PilQ (33), adopts a similar β structure to GspC, a component of the T2SS that has a related function in binding to its cognate secretin GspD (34). Moreover, there are also similarities in the ways the two secretin-binding proteins recognize the secretin α/β domains in PilQ and GspD (8,34). The PilP lipoprotein is formed from a long, flexible N terminus, which contains the lipid attachment site, and a globular C-terminal domain (35).…”

On the path to uncover the bacterial type II secretion system

“…Moreover, while results obtained in K. oxytoca and V. cholerae [42][43][44] with GFP-fused Gsp proteins indicate a circumferential distribution of the machinery into foci, the P. aeruginosa Xcp secreton was proposed to be polar. This was shown by adding a Lumino tag onto XcpR or XcpS or by the visualization of protease secretion with an intramolecularly quenched casein conjugate [45].…”

“…GspC P was shown to be active as a dimer and self-associates by its TM domain, which is not a simple membrane anchor but plays an active role in the function of the protein [72]. The HR domain of V. cholerae GspC P (EpsC P ) was shown to interact directly with the periplasmic N 0 domain of the secretin EpsD Q [44,73]. This interaction was also seen in E. chrysanthemi where the interaction site of OutC P on OutD Q was localized between residues 139 and 158 of the HR domain.…”