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Purification, crystallization, X‐ray diffraction analysis and phasing of an engineered single‐chain PvuII restriction endonuclease

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The restriction endonuclease PvuII from Proteus vulgaris has been converted from its wild‐type homodimeric form into the enzymatically active single‐chain variant scPvuII by tandemly joining the two subunits through the peptide linker Gly‐Ser‐Gly‐Gly. scPvuII, which is suitable for the development of programmed restriction endonucleases for highly specific DNA cleavage, was purified and crystallized. The crystals diffract to a resolution of 2.35 Å and belong to space group P42, with unit‐cell parameters a = b = 101.92, c = 100.28 Å and two molecules per asymmetric unit. Phasing was successfully performed by molecular replacement.
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Keywords: PvuII; programmed restriction endonucleases; scPvuII

Document Type: Research Article

Publication date: October 1, 2007

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