Diverse substrate recognition and hydrolysis mechanisms of human NUDT5

Arimori, Takao; Tamaoki, Haruhiko*; Nakamura, Teruya*; Kamiya, Hiroyuki*; Ikemizu, Shinji*; Takagi, Yasumitsu*; Ishibashi, Toru*; Harashima, Hideyoshi*; Sekiguchi, Mutsuo*; Yamagata, Yuriko*

Human NUDT5 (hNUDT5) hydrolyzes various modified nucleoside diphosphates including 8-oxo-dGDP, 8-oxo-dADP and ADP-ribose (ADPR). However, the structural basis of the broad substrate specificity remains unknown. Here, we report the crystal structures of hNUDT5 complexed with 8-oxo-dGDP and 8-oxo-dADP. These structures reveal an unusually different substrate-binding mode. In particular, the positions of two phosphates ( and phosphates) of substrate in the 8-oxo-dGDP and 8-oxo-dADP complexes are completely inverted compared with those in the previously reported hNUDT5-ADPR complex structure. This result suggests that the nucleophilic substitution sites of the substrates involved in hydrolysis reactions differ despite the similarities in the chemical structures of the substrates and products. To clarify this hypothesis, we employed the isotope-labeling method and revealed that 8-oxo-dGDP is attacked by nucleophilic water at P, whereas ADPR is attacked at P.