The Vibrio vulni®cus nuclease, Vvn, is a non-speci®c periplasmic nuclease capable of digesting DNA and RNA. The crystal structure of Vvn and that of Vvn mutant H80A in complex with DNA were resolved at 2.3 A Ê resolution. Vvn has a novel mixed a/b topology containing four disul®de bridges, suggesting that Vvn is not active under reducing conditions in the cytoplasm. The overall structure of Vvn shows no similarity to other endonucleases; however, a known`bba± metal' motif is identi®ed in the central cleft region. The crystal structure of the mutant Vvn±DNA complex demonstrates that Vvn binds mainly at the minor groove of DNA, resulting in duplex bending towards the major groove by~20°. Only the DNA phosphate backbones make hydrogen bonds with Vvn, suggesting a structural basis for its sequence-independent recognition of DNA and RNA. Based on the enzyme±sub-strate and enzyme±product structures observed in the mutant Vvn±DNA crystals, a catalytic mechanism is proposed. This structural study suggests that Vvn hydrolyzes DNA by a general single-metal ion mechanism, and indicates how non-speci®c DNA-binding proteins may recognize DNA.

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