Townson Sharon A, Samuelson James C, Vanamee Eva Scheuring, Edwards Thomas A, Escalante Carlos R, Xu Shuang-Yong, Aggarwal Aneel K
Structural Biology Program, Department of Physiology and Biophysics, Mount Sinai School of Medicine, 1425 Madison Avenue, New York, NY 10029, USA.
J Mol Biol. 2004 May 7;338(4):725-33. doi: 10.1016/j.jmb.2004.02.074.
We report here the structure of BstYI, an "intermediate" type II restriction endonuclease with overlapping sequence specificities to BamHI and BglII. BstYI, a thermophilic endonuclease, recognizes and cleaves the degenerate hexanucleotide sequence 5'-RGATCY-3' (where R=A or G and Y=C or T), cleaving DNA after the 5'-R on each strand to produce four-base (5') staggered ends. The crystal structure of free BstYI was solved at 1.85A resolution by multi-wavelength anomalous dispersion (MAD) phasing. Comparison with BamHI and BglII reveals a strong structural consensus between all three enzymes mapping to the alpha/beta core domain and residues involved in catalysis. Unexpectedly, BstYI also contains an additional "arm" substructure outside of the core protein, which enables the enzyme to adopt a more compact, intertwined dimer structure compared with BamHI and BglII. This arm substructure may underlie the thermostability of BstYI. We identify putative DNA recognition residues and speculate as to how this enzyme achieves a "relaxed" DNA specificity.
我们在此报道了BstYI的结构，它是一种“中间型”II型限制性内切酶，对BamHI和BglII具有重叠的序列特异性。BstYI是一种嗜热内切酶，识别并切割简并六核苷酸序列5'-RGATCY-3'（其中R = A或G，Y = C或T），在每条链上5'-R之后切割DNA，产生四碱基（5'）交错末端。通过多波长反常散射（MAD）相位法在1.85Å分辨率下解析了游离BstYI的晶体结构。与BamHI和BglII的比较揭示了这三种酶在映射到α/β核心结构域和参与催化的残基之间存在很强的结构一致性。出乎意料的是，BstYI在核心蛋白之外还包含一个额外的“臂”亚结构，与BamHI和BglII相比，这使得该酶能够采用更紧凑、相互缠绕的二聚体结构。这个臂亚结构可能是BstYI热稳定性的基础。我们确定了推定的DNA识别残基，并推测了这种酶如何实现“宽松”的DNA特异性。
