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| The EMBO Journal
(2000) 19, 758–766, doi:10.1093/emboj/19.4.758 |
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| DNA bending and a flip-out mechanism for base excision by the helix–hairpin–helix DNA glycosylase, Escherichia coli AlkA |
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| Thomas Hollis1, Yoshitaka Ichikawa2 and Tom Ellenberger1 |
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1 Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, 240 Longwood Avenue, Boston, MA 02115, USA 2 Department of Pharmacology and Medical Sciences, Johns Hopkins University School of Medicine, Baltimore, MD 21205, USA To whom correspondence should be addressed Tom Ellenberger, tome@hms.harvard.edu Received 1 November 1999; Revised 6 December 1999; Accepted 6 December 1999. |
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| Abstract |
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| The Escherichia coli AlkA protein is a base excision repair glycosylase that removes a variety of alkylated bases from DNA. The 2.5 Å crystal structure of AlkA complexed to DNA shows a large distortion in the bound DNA. The enzyme flips a 1-azaribose abasic nucleotide out of DNA and induces a 66° bend in the DNA with a marked widening of the minor groove. The position of the 1-azaribose in the enzyme active site suggests an SN1-type mechanism for the glycosylase reaction, in which the essential catalytic Asp238 provides direct assistance for base removal. Catalytic selectivity might result from the enhanced stacking of positively charged, alkylated bases against the aromatic side chain of Trp272 in conjunction with the relative ease of cleaving the weakened glycosylic bond of these modified nucleotides. The structure of the AlkA–DNA complex offers the first glimpse of a helix–hairpin–helix (HhH) glycosylase complexed to DNA. Modeling studies suggest that other HhH glycosylases can bind to DNA in a similar manner. |
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| Keywords: AlkA, glycosidase mechanism, helix–hairpin–helix, 3-methyladenine DNA glycosylase, protein–DNA complex |
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