The disordered C-tail of SecB modulates holdase activity and translocase engagement

Secretory chaperones enable preproteins that are secreted via the Sec post-translational pathway, to maintain their unfolded, translocation-competent states. SecB, a proteobacterial secretory chaperone, has a short, disordered C-terminal tail whose function remains largely unexplored. Here, we investigate the role of this tail using a combination of in vivo and in vitro functional assays, single-molecule FRET, hydrogen-deuterium exchange mass spectrometry, and isothermal titration calorimetry. We show that the C-terminal tail serves two functions in SecB: it reinforces holdase activity by prolonging client retention in an expanded, translocation-competent conformation, and it attenuates SecA binding affinity by maintaining the β-sheet docking surface in a dynamic state. The tail-deletion phenotype partially mirrors the dynamic signature of client-loaded SecB, suggesting the tail acts as a conditional element coupling client occupancy to SecA engagement. These findings identify the disordered C-terminal tail as a bifunctional regulator of client retention and translocase handoff in the Sec pathway.