The crystal structure of the DNA‐binding domain of E. coli SSB (EcoSSB) has been determined to a resolution of 2.5 Å. This is the first reported structure of a prokaryotic SSB. The structure of the DNA‐binding domain of the E. coli protein is compared to that of the human mitochondrial SSB (HsmtSSB). In spite of the relatively low sequence identity between them, the two proteins display a high degree of structural similarity. EcoSSB crystallises with two dimers in the asymmetric unit, unlike HsmtSSB which contains only a dimer. This is probably a consequence of the different polypeptide chain lengths in the EcoSSB heterotetramer. Crucial differences in the dimer‐dimer interface of EcoSSB may account for the inability of EcoSSB and HsmtSSB to form cross‐species heterotetramers, in contrast to many bacterial SSBs.
The crystal structure of the dodecanucleotide d(CGCAAGCTGGCG) has been determined to a resolution of 2.5 A and refined to an R factor of 19.3% for 1710 reflections.The sequence crystallizes as a B-type double helix, with two G(ant)-A(syn) base pairs. These are stabilized by three-center hydrogen bonds to pyrimidines that induce perturbations in base-pair geometry. The central AGCT region of the helix has a wide (>6 A) minor groove.Mispairing in DNA can occur through genetic recombination events or as errors in replication (1). These mismatches have mutational consequences unless they are corrected by repair processes such as excision repair (2), which in Escherichia coli involves the uvrD gene product DNA helicase II and the dam methylase (3, 4). The A-G mismatch, which results in transversions, appears to have specific excision repair mechanisms associated with it, at least in E. coli (2), with the MutT protein appearing to prevent G-A pairing during replication (5). A-G repairs are less efficiently repaired in some mammalian cells than other heterogeneous mismatches, possibly because of structural differences between them (6).A number of distinct base-pairing possibilities have been suggested for A-G pairing (7,8), including the involvement of imino tautomers for either adenine or guanine. NMR spectroscopy has shown that G(anti)-A(anti) mismatches occur in some oligonucleotides (9, 10) with sequences around the mismatches of 5'-R-A(=G)-G(=A)-Y-3' and 5'-R-A(=G)-R-3', respectively (where = donates a mispair, R is a purine, and Y is a pyrimidine). Alternative G(syn)-A(anti) mispairing has been found at low pH by NMR methods in oligonucleotides with A(=G) flanked at both 5' and 3' sides by either guanine or cytosine (11). X-ray crystallographic studies have revealed both of these possibilities, as well as the less expected G(anti)-A(syn) arrangement (12, 13), in the sequence d(CGC-GAATTAGCG). The G(anti)-A(anti) pairing has been found for the two contiguous A-G base pairs in the crystal structure of the sequence d(CCAAGCTTGG) (14, 15) and G(syn)-A(anti) in the dodecanucleotide d(CGCAAATTG-GCG) (16). The latter structure has by implication protonation at N1 of the adenine to achieve the observed base pairing. Intensity data were collected on an Enraf-Nonius FAST area detector at Birkbeck College for a crystal of size 0.5 x 0.06 x 0.02 mm mounted in a sealed capillary tube. A total of 9842 reflections were collected to a resolution of 2.2 A at 70C. Only the 2495 reflections to 2.5 A (with a merging R factor of 11.9%) were used in the analysis since the merging agreement factor for the (mostly weak) reflections in the range of 2.2-2.5 A was unacceptably high. A total of 1710 unique reflections with F0> 3or (F'0) were obtained.
5' CGCAAGCTGGCGThe structure was solved by molecular replacement using the established structure of the dodecamer d(CGCGAAT-TCGCG) (17). The initial stages ofrefinement were undertaken using the CORELS constrained-restrained refinement package (18). The coordinates of the Dickerson dod...
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.