Adapters drive context-dependent Mycobacterial ClpX activity by altering conformational sampling
The AAA+ ATPase ClpX recognizes diverse substrates and plays a crucial role in cellular homeostasis. Mycobacterium tuberculosis (Mtb) ClpX binds to two structurally distinct adaptors- the Single-stranded DNA binding protein (SSB) and the Small protein B (SmpB).  The mechanistic features of adaptor-dependent modulation of ClpX activity and its functional implications still remain poorly understood.  Results show that both adaptors bind to the N-terminal domain of ClpX, albeit with different stoichiometry. While SSB binding enhances ATPase activity, SmpB does not.  Structural and biochemical experiments revealed that both adaptors substantially reduce conformational sampling and drive ClpX into a more compact, homogenous quaternary arrangement.  The interactome of the adaptors and ClpX indicate features that distinguish adaptor-dependent from adaptor-independent ClpX activity.  Changes in the expression profile in Mtb cells upon over-expression of either SSB or SmpB suggest functional linkages between the protein recycling mechanism and transcription.  Mtb ClpX is thus both- an unfoldase driving protein maturation and a checkpoint regulator synchronizing transcription and translational processes in this slow-growing human pathogen. Overall design: To investigate the downstream role of SSB and SmpB, the transcriptomics of the vector control Rv strain (Rv::pN), and overexpressing strains of ssb (Rv::pN-ssb) and smpB (Rv::pN-smpB) was performed