Tyrosylprotein sulfotransferases (TPSTs) catalyze O-sulfation of tyrosine residues on secreted and membrane proteins, but the molecular basis for their stimulation by metal ions remains unclear. We determined the structures of the catalytic domain of human TPST2 with PAP and Na+ (1.75 Å) or Mn2+ (2.00 Å) bound and identified two conserved octahedral metal-binding sites. Anomalous diffraction at metal absorption edges confirmed the identity of the bound metals and demonstrated specific Mn2+ binding. The Na+- and Mn2+-bound structures closely superimposed, suggesting activation without large conformational changes. Structural comparison with the apo structure and ensemble refinement revealed differences in local dynamics around the metal binding sites. The flexible α3-helix and α12-α13 loop in the apo structure were stabilized by Na+ binding and further rigidified by Mn2+ binding. These findings support an activation-by-ordering mechanism in which Na+ binding generates a pre-activated state, with Mn2+ subsequently establishing a catalytically competent ordering that lowers the entropic barrier at the active-site entrance. This framework reconciles longstanding biochemical observations and suggests that Mn2+ availability within the Golgi can tune TPST2-dependent signaling.
Jin et al. (Fri,) studied this question.