Solution structure of the C-terminal single-stranded DNA-binding domain of Escherichia coli topoisomerase I

• Published in: Biochemistry (Easton) 1995-06, Vol.34 (23), p.7622-7628
• Main Authors: Yu, Liping, Zhu, Chang-Xi, Tse-Dinh, Yuk-Ching, Fesik, Stephen W
• Format: Article
• Language: English
• Subjects: Bacterial Proteins - chemistry; Base Sequence; Binding Sites; DNA Topoisomerases, Type I - chemistry; DNA, Single-Stranded - metabolism; DNA-Binding Proteins - chemistry; Escherichia coli; Escherichia coli - enzymology; Models, Molecular; Molecular Sequence Data; Oligodeoxyribonucleotides - metabolism; Peptide Fragments - chemistry; Protein Structure, Tertiary; Solutions
• ISSN: 0006-2960; 1520-4995
• DOI: 10.1021/bi00023a008

Escherichia coli DNA topoisomerase I catalyzes the interconversion of different topological forms of DNA. In this paper we describe NMR studies of a 14K C-terminal fragment of this enzyme that binds preferentially to single-stranded DNA and enhances the enzyme's ability to relax negatively supercoiled DNA under high salt conditions. The 1H, 13C, and 15N resonances of the protein were assigned from a number of heteronuclear multidimensional NMR experiments, and the three-dimensional structure of the protein was determined from a total of 2188 NMR-derived restraints. The root-mean-square deviation about the mean coordinate positions for residues 13-120 is 0.68 +/- 0.11 A for the backbone atoms and 1.09 +/- 0.09 A for all heavy atoms. The overall fold, which consists of two four-stranded beta-sheets separated by two helices, differs from other DNA- and RNA-binding proteins such as gene 5, cold shock protein, and hnRNP C. From an analysis of the changes in chemical shift upon the addition of single-stranded DNA, the location of the oligonucleotide binding site was determined. The binding site consists of a beta-sheet containing positively charged and aromatic amino acids and, in spite of its different structure, is similar to that found in other proteins that bind single-stranded oligonucleotides.