Selenium-assisted nucleic acid crystallography: Use of phosphoroselenoates for MAD phasing of a DNA structure

Christopher J. Wilds, Rekha Pattanayek, Chongle Pan, Zdzislaw Wawrzak, Martin Egli*

*Corresponding author for this work

    Research output: Contribution to journalArticlepeer-review

    79 Scopus citations

    Abstract

    The combination of synchrotron radiation and a variety of atoms or ions (either covalently attached to the biomolecule prior to crystallization or soaked into crystals) that serve as anomalous scatterers constitutes a powerful tool in the X-ray crystallographer's repertoire of structure determination techniques. Phosphoroselenoates in which one of the nonbridging phosphate oxygens in the backbone is replaced by selenium offer a simplified means for introducing an anomalous scatterer into oligonucleotides by conventional solid-phase synthesis. Unlike other methods that are used to derivatize DNA or RNA by covalent attachment of a heavy atom (i.e., bromine at the C5 position of pyrimidines), tedious synthesis of specialized nucleosides is not required. Introduction of selenium is readily accomplished in solid-phase oligonucleotide synthesis by replacing the standard oxidation agent with a solution of potassium selenocyanide. This results in a diastereomeric mixture of phosphoroselenoates that can be separated by strong anion-exchange HPLC. As a test case, all 10 DNA hexamers of the sequence CGCGCG containing a single phosphoroselenoate linkage (PSe) were prepared. Crystals were grown for a subset of them, and the structure of [d(CPSe-GCGCG)]2 was determined by the multiwavelength anomalous dispersion technique and refined to 1.1 Å resolution.

    Original languageEnglish (US)
    Pages (from-to)14910-14916
    Number of pages7
    JournalJournal of the American Chemical Society
    Volume124
    Issue number50
    DOIs
    StatePublished - Dec 18 2002

    ASJC Scopus subject areas

    • Catalysis
    • Chemistry(all)
    • Biochemistry
    • Colloid and Surface Chemistry

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