General scheme of the reaction: Our study does not deviate from the general scheme of two metal ion catalysis. RNase H catalyzed the cleavage of the RNA strand of the DNA/RNA hybrid via two catalytic magnesium ions. The nucleophilic water (WATnuc) was bound to MgA and oriented for nucleophilic attack on the phosphate. The O3' leaving group was bound to MgB (Figure ). The reaction proceeded via formation of a pentavalent phosphorane transition state (TS1, Figure ) after the WATnuc is deprotonated, a high energy intermediate state (INT, Figure ) and a second transition state (TS2, Figure ), where the leaving group is protonated.