Figure 2.

Geometry and flexibility of catalytic residues in the active sites of CCR4 and CAF1. (A) Active site of Hs CCR4a with the conserved side chains of the EEP nuclease family shown as green sticks. Other important side chains are also shown as sticks. Residues mutated in the course of this work are labeled in magenta. Superposition with the active site of Hs CCR4b (black, PDB-ID 3ngn) (29) demonstrates the conservation of sequence and structure apart from the HWDP loop and the loop L(β11–β12), which appear to be flexible elements (red). (B) Active site of Hs CAF1a with the conserved side chains of the DEDDh nuclease family shown as green sticks. Other important side chains are also shown as sticks. Residues mutated in the course of this work are labeled in magenta. Superposition with a previously determined structure of Hs CAF1a (black, PDB-ID 4gmj) (48) indicates malleability of H157 and H225 and shows the position of the C-terminal peptide (red), which may interfere with the access of an RNA substrate. (C) Active site of Hs CCR4a including a crude model of an RNA 3′-terminal end. The 3′-terminal adenine corresponds to the AMP molecule found in the structure of Hs CCR4b (PDB-ID 3ngn) (29). Modelling of the penultimate adenine was guided by a DNA ligand of Hs CCR4b (PDB-ID 3ngo) (29), followed by energy minimization. (D) Active site of Hs CAF1a including a crude model of an RNA 3′-terminal end. The two 3′-terminal adenines were placed according to a structure of the PARN 3′-deadenylase in complex with RNA (PDB-ID 2a1r) (20). Oxygen atoms, red; nitrogen atoms, blue; magnesium ions, magenta.