VadK, a Non-Canonical Kinase that Regulates the Methylcitrate Cycle and is Essential for Mycobacterium tuberculosis Virulence.

The evolution of new enzymatic functions is constrained and guided by the architecture of an organism's metabolic and regulatory networks as well as by environmental constraints. Here, we identify a previously uncharacterized kinase that has evolved from pyruvate phosphate dikinase (PPDK). Through biochemical and systems-level analyses, we show that this enzyme, encoded by Rv1127c in Mycobacterium tuberculosis (Mtb), has diverged from its ancestral role in central carbon metabolism to function as a histidine kinase in pathogenic mycobacteria and related species. We designate this enzyme Virulence Associated DiKinase (VadK), reflecting its ability to autophosphorylate and role in coordinating metabolism and virulence. VadK is essential for the utilization of exogenous carbon sources critical for survival within the host and is required for Mtb pathogenicity in murine models of tuberculosis. Furthermore, VadK interacts with key enzymes of the methylcitrate cycle, and 13C-metabolic flux analysis indicates that it fine-tunes flux through this pathway, with elevated flux proving growth limiting. Together, these findings identify VadK as a previously unrecognized regulatory kinase that integrates metabolic control with virulence in Mtb, revealing a new facet of metabolic regulation in bacterial pathogenesis and a potential target for therapeutic intervention.