Structural insights into the mechanism of Escherichia coli YmdB: A 2'-O-acetyl-ADP-ribose deacetylase

Abstract The Escherichia coli protein YmdB belongs to the macrodomain protein family, which can bind ADP-ribose (ADPr) and its derivatives. Recently, YmdB was reported to be capable of deacetylating O -acetyl-ADP-ribose ( O AADPr) to yield ADPr and free acetate. To study the substrate specificity and catalytic mechanism, the crystal structures of E. coli YmdB in complex with ADPr, double mutant N25AD35A complexed with 2′- O AADPr, and Y126A/ADPr complex were solved at 1.8 A, 2.8 A and 3.0 A resolution, respectively. Structural and biochemical studies reveal that YmdB has substrate specificity against 2′- O AADPr. The conserved residues Asn25 and Asp35 are crucial for catalytic activity, and an active water molecule is proposed as the nucleophile to attack the acetyl group of 2′- O AADPr. Our findings indicate that the conserved phenyl group of Tyr126 plays a crucial role in catalytic activity by stabilizing the right orientation of distal ribose and that Gly32 may be important for activity by interacting with the acetyl group of 2′- O AADPr. Based on these observations, a model of YmdB in complex with 2′- O AADPr was made to illustrate the proposed catalytic mechanism of YmdB.