Functional and mechanistic insights into the stealth protein full-length CpsY is conducive to understanding immune evasion mechanisms by Mycobacterium tuberculosis.

Mycobacterium tuberculosis (Mtb) is a crucial and destructive intracellular pathogen responsible for causing tuberculosis (TB), a disease of substantial morbidity and mortality. Mtb capsular polysaccharides can misdirect the host's immune response pathways, resulting in additional challenges in TB treatment. These capsule polysaccharides are biosynthesized by a series of stealth proteins including CpsY. Our prior investigations elucidated the structural and functional information of the central domain (aa 201-520) of CpsY within Mtb. However, within the host milieu, it is the full-length iteration of CpsY, rather than its truncated form CpsY201-520, that assumes pivotal roles in immune evasion. Consequently, investigating the functional mechanism of full-length CpsY is extremely important. Here, we found that the indispensable role of four conserved regions (CR1-CR4) in governing the phosphotransferase activity of full-length CpsY. Notably, the deletion of S2 (ΔS2) dramatically increased the activity compared to the wild-type (WT) full-length CpsY, thereby revealing S2 in the regulatory dynamics governing the inactivation and activation of full-length CpsY. The gene cpsY helps Mtb to survive in macrophages. Our findings were useful for the development of vaccines and immunotherapies targeting Mtb.