In this paper we have examined the mechanism of tyrosine O-sulfonation catalyzed by human TPST-2. Our computations, in agreement with Teramoto's hypothesis, indicate a concerted SN2-like reaction (with an activation barrier of 18.2 kcal mol(-1)) where the tyrosine oxygen is deprotonated by Glu(99) (base catalyst) and simultaneously attacks as a nucleophile the sulfuryl group. For the first time, using a quantum mechanics protocol of alanine scanning, we identified unequivocally the role of the amino acids involved in the catalysis. Arg(78) acts as a shuttle that "assists" the sulfuryl group moving from the 3'-phosphoadenosine-5'-phosphosulfate molecule to threonine and stabilizes the transition state (TS) by electrostatic interactions. The residue Lys(158) keeps close the residues participating in the overall H-bond network, while Ser(285), Thr(81), and Thr(82) stabilize the TS via strong hydrogen interactions and contribute to lower the activation barrier.