4OC8

DNA modification-dependent restriction endonuclease AspBHI

Summary for 4OC8

• Entry DOI: 10.2210/pdb4oc8/pdb
• Related: 4F0Q
• Descriptor: restriction endonuclease AspBHI, PHOSPHATE ION (3 entities in total)
• Functional Keywords: dna cleavage, dna restriction enzymes, dna-binding proteins, tetramerization, models, molecular, azoarcus, protein multimerization, protein structure, tertiary, hydrolase
• Biological source: Azoarcus sp.
• Total number of polymer chains: 4
• Total formula weight: 172022.10

Authors

Horton, J.R. (deposition date: 2014-01-08, release date: 2014-03-19, Last modification date: 2024-02-28)

Primary citation

Horton, J.R.,Nugent, R.L.,Li, A.,Mabuchi, M.Y.,Fomenkov, A.,Cohen-Karni, D.,Griggs, R.M.,Zhang, X.,Wilson, G.G.,Zheng, Y.,Xu, S.Y.,Cheng, X.
Structure and mutagenesis of the DNA modification-dependent restriction endonuclease AspBHI.
Sci Rep, 4:4246-4246, 2014

PubMed Abstract: The modification-dependent restriction endonuclease AspBHI recognizes 5-methylcytosine (5mC) in the double-strand DNA sequence context of (C/T)(C/G)(5mC)N(C/G) (N = any nucleotide) and cleaves the two strands a fixed distance (N12/N16) 3' to the modified cytosine. We determined the crystal structure of the homo-tetrameric AspBHI. Each subunit of the protein comprises two domains: an N-terminal DNA-recognition domain and a C-terminal DNA cleavage domain. The N-terminal domain is structurally similar to the eukaryotic SET and RING-associated (SRA) domain, which is known to bind to a hemi-methylated CpG dinucleotide. The C-terminal domain is structurally similar to classic Type II restriction enzymes and contains the endonuclease catalytic-site motif of DX20EAK. To understand how specific amino acids affect AspBHI recognition preference, we generated a homology model of the AspBHI-DNA complex, and probed the importance of individual amino acids by mutagenesis. Ser41 and Arg42 are predicted to be located in the DNA minor groove 5' to the modified cytosine. Substitution of Ser41 with alanine (S41A) and cysteine (S41C) resulted in mutants with altered cleavage activity. All 19 Arg42 variants resulted in loss of endonuclease activity.

PubMed: 24604015
DOI: 10.1038/srep04246

Experimental method

X-RAY DIFFRACTION (2.884 Å)