Abstract:

<p>A recently identified and widely prevalent prokaryal gene cluster encodes a suite of enzymes with imputed roles in nucleic acid repair. The enzymes are: MPE, a DNA endonuclease; Lhr-Core, a 3&#39;-5&#39; DNA helicase; LIG, an ATP-dependent DNA ligase; and Exo, a metallo-β-lactamase-family nuclease. Bacterial and archaeal MPE proteins belong to the binuclear metallophosphoesterase superfamily that includes the well-studied DNA repair nucleases Mre11 and SbcD. Here we report that the MPE protein is a manganese-dependent DNA endonuclease that incises either linear single strands or the single-strand loops of stem-loop DNA structures. MPE has feeble activity on duplex DNA. A crystal structure of MPE at 2.2 Å&nbsp;resolution revealed that the active site includes two octahedrally coordinated manganese ions. Seven signature amino acids of the binuclear metallophosphoesterase superfamily serve as the enzymic metal ligands in MPE: Asp33, His35, Asp78, Asn112, His124, His146, and His158. A swath of positive surface potential on either side of the active site pocket suggests a binding site for the single-strand DNA substrate. The structure of MPE differs significantly from Mre11 and SbcD in several key respects: (i) MPE is a monomer, whereas Mre11 and SbcD are homodimers; (ii) MPE lacks the capping domain present in Mre11 and SbcD; and (iii) the topology of the bsandwich that comprises the core of the metallophosphoesterase fold differs in MPE Mre11 and SbcD. We surmise that MPE exemplifies a novel clade of DNA endonuclease within the binuclear metallophosphoesterase superfamily.</p>