Figure 1.

Illustration of the active site interactions in HHR and its self-cleavage mechanism. The N1 position of guanine in the putative general base G12 (blue) needs to be deprotonated before acting as a proton acceptor to deprotonate the 2′-OH in C17 (red), which will then act as the nucleophile to attack the phosphorous. Recent studies(15, 16) indicate that there could be a Mg²⁺ directly bound at the Hoogsteen face of G12 (“G-site”) to facilitate its deprotonation. Another Mg²⁺ is believed to play the role of activating the 2′-OH of the general acid G8 (green) by migrating from the binding site at N7 of G10.1 (“C-site”) observed crystallographically into a bridging position (“B-site”) with the scissile phosphate, in accord with thio/rescue effect experiments(29, 39, 57). In this bridging position, the Mg²⁺ can coordinate the 2′-OH of G8, increasing its acidity, and facilitating proton transfer to the O5′ leaving group in the general acid step of the reaction(8).