Structure of HhaI endonuclease with cognate DNA at an atomic resolution of 1.0 Å

Journal Article · 27 December 2019 · Nucleic Acids Research

DOI: https://doi.org/10.1093/nar/gkz1195 · OSTI ID:1603149

Horton, John R. ^[1]; Yang, Jie ^[1]; Zhang, Xing ^[1]; Petronzio, Theresa ^[2]; Fomenkov, Alexey ^[2]; Wilson, Geoffrey G. ^[2]; Roberts, Richard J. ^[2]; Cheng, Xiaodong  ^[1]

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1. Univ. of Texas, Houston, TX (United States). Anderson Cancer Center
2. New England Biolabs, Inc., Ipswich, MA (United States)

HhaI, a Type II restriction endonuclease, recognizes the symmetric sequence 5'-GCG↓C-3' in duplex DNA and cleaves (‘↓’) to produce fragments with 2-base, 3'-overhangs. We determined the structure of HhaI in complex with cognate DNA at an ultra-high atomic resolution of 1.0 Å. Most restriction enzymes act as dimers with two catalytic sites, and cleave the two strands of duplex DNA simultaneously, in a single binding event. HhaI, in contrast, acts as a monomer with only one catalytic site, and cleaves the DNA strands sequentially, one after the other. HhaI comprises three domains, each consisting of a mixed five-stranded β sheet with a defined function. The first domain contains the catalytic-site; the second contains residues for sequence recognition; and the third contributes to non-specific DNA binding. The active-site belongs to the ‘PD-D/EXK’ superfamily of nucleases and contains the motif SD-X11-EAK. The first two domains are similar in structure to two other monomeric restriction enzymes, HinP1I (G↓CGC) and MspI (C↓CGG), which produce fragments with 5'-overhangs. The third domain, present only in HhaI, shifts the positions of the recognition residues relative to the catalytic site enabling this enzyme to cleave the recognition sequence at a different position. The structure of M.HhaI, the biological methyltransferase partner of HhaI, was determined earlier. Together, these two structures represent the first natural pair of restriction-modification enzymes to be characterized in atomic detail.

• Research Organization: Argonne National Laboratory (ANL), Argonne, IL (United States)
• Sponsoring Organization: USDOE; National Institutes of Health (NIH)
• OSTI ID: 1603149
• Journal Information: Nucleic Acids Research, Vol. 48, Issue 3; ISSN 0305-1048
• Publisher: Oxford University PressCopyright Statement
• Country of Publication: United States
• Language: ENGLISH