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The following HTML text is provided to enhance online readability. Many aspects of typography translate only awkwardly to HTML. Please use the page image as the authoritative form to ensure accuracy. Table 1. Comparison of NOEs derived from various forms of DNA
double-stranded DNA switch to new base pairs during heteroduplex formation (base pair switch), are still unanswered questions in homologous recombination. Two different mechanisms have been considered. In the first mechanism, RecA disrupts the base pairs of double-stranded DNA and promotes an annealing reaction between single-stranded DNA and a strand of double-stranded DNA. In the other mechanism, the recognition of sequence homology occurs through non-Watson-Crick interactions without the need for the disruption of the base pairs of the double-stranded DNA, such as interactions that form a triplex. Various and extensive attempts, such as chemical or enzymatic probing experiments, have been carried out to elucidate the molecular mechanism of heteroduplex joint formation by RecA (see refs. 55–58 for review), but these studies have not succeeded in giving a unified view of the mechanism involved. For example, whereas many studies have supported the idea that an incoming single-stranded DNA interacts with double-stranded DNA in its minor groove for the recognition of sequence homology by RecA (59–63), results contradicting this were published recently (64). The Three-Dimensional Molecular Structure of Single-Stranded Oligo-DNA Bound to RecAIn an attempt to understand the mechanisms of homologous pairing and strand exchange, we analyzed the three-dimensional molecular structure of single-stranded oligo-DNA bound to RecA in the presence of an ATP analogue, ATPγS, by use of the transferred nuclear Overhauser effect (NOE) method, a technique of NMR spectroscopy. This NMR technique is frequently used for the structural analysis of small ligands bound to proteins (65, 66). We used single-stranded oligo-DNA with 3–6 bases and some variations in sequence. The NOEs obtained are clearly different from those of either B-form or A-form DNA (ref. 67 and T.N., unpublished observations; Table 1). The structure calculations applying a standard simulated annealing protocol gave a unique well-defined extended DNA structure for each single-stranded oligo-DNA (67). When RecA was replaced by Saccharomyces cerevisiae Rad51, the spectra were very similar to those obtained with RecA, indicating that RecA and Rad51 induce a common extended structure in single-stranded DNA upon binding (68). The results from the analysis (Fig. 1; Protein Data Bank ID 3REC; refs. 67 and 68) revealed that (i) the extended single- 
Fig. 1. A model for the extended single-stranded DNA structure induced by binding of RecA/Rad51. (A) The model structure and summary of NOE intensities. (B and C) Comparison of the model structure of the extended single-stranded DNA (B) with a part of B-form DNA (C). The extended single-stranded DNA structure, in which the distance between bases is ca. 5 Å, contains hydrophobic 2′ methylene-base interactions, instead of the base-base stacking found in B-form DNA (the distance between bases: 3.4 Å). [Reproduced with permission from ref. 67 (Copyright 1997, National Academy of Sciences).] stranded DNA structure contains hydrophobic deoxyribose-base stacking in which the 2′ methylene moiety of each deoxyribose is placed above the base of the following residue, instead of the base-base stacking found in B-form DNA; (ii) the distance between neighboring bases is expanded to about 5 Å from the 3.4 Å of B-form DNA; (iii) the structure is specific to DNA, because the 2′ methylene-base interaction (the interaction of the 2′ methylene moiety with the aromatic ring of the next base) is likely to be a CH/π interaction, a weak attractive molecular force occurring between the CH groups and π-systems observed in various biomacromolecules, which would contribute to the stabilization of the extended structure; and (iv) the extended structure requires RecA or Rad51 and (at least in the case of RecA) an ATP analogue (69). To understand the roles of the proteins that are required for the induction and maintenance of the extended DNA structure, structural information about the DNA binding sites on the proteins is required, but was not obtained in this study, because of a technical limit of transferred NOE method. The distance between bases by itself explains the well-known 1.5-fold longer length of single-stranded DNA as compared with B-form double-stranded DNA with the same sequence (48), because the bases have been shown to be almost perpendicular
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