Type III secretion of transmembrane proteins and the case of the Salmonella´s chaperone, SscB

Type III secretion (T3S) systems are present in Gram-negative bacteria and consist of a needle-like structure that creates a direct channel for the delivery of bacterial proteins into host cells, ultimately promoting bacterial survival. Among the proteins delivered are those containing transmembrane domains (TMD). These can interact with a T3S chaperone (T3SC), which protects the more hydrophobic TMDs from erroneous recognition and targeting to bacterial membranes. This interaction should be critical for an efficient and correct T3S. Here, we aimed to unveil the key features and processes that underlie the T3S of TMD-effectors.

In Salmonella, SscB/SseF are a chaperone/TMD-effector pair. Biophysical characterization of purified SscB and analysis of the predicted structure showed that SscB is structurally similar to T3SCs that bind translocators (TMD-proteins). Interestingly, a consensus amino acid sequence, which is conserved on the translocators of Salmonella and other bacterial species, was identified in SseF but also in the TMD-effector SseG. SscB interacts and promotes secretion of SseF, as seen previously, but also can interact and aid the secretion of SseG. We showed that mutagenesis of the consensus amino acid sequence abolished T3S of SseF and SseG. Next, it was observed that SscB and SseF co-stabilized but SseG was stable without the presence of the chaperone. This suggests that efficient SscB/SseF interaction is more critical than that of SscB/SseG. Moreover, since these three proteins are encoded adjacently in an operon, sscB-sseF-sseG, we have analyzed the effects of changing gene order on T3S. A decrease in the T3S of SseF was observed when sscB was relocated downstream of sseF.

Overall, SscB shares features with chaperones of translocators, which challenges the current classification of T3SC. This suggests that the structure of the T3SC depends on the structural characteristics of their interacting partners rather than their timing of secretion. The gene order is important for efficient targeting to T3S, by possibly allowing for a more rapid interaction of SscB/SseF, which leads to protein co-stabilization and protection of the TMD of SseF.