CRYSTAL-STRUCTURE OF A REPLICATION FORK SINGLE-STRANDED-DNA BINDING-PROTEIN (T4 GP32) COMPLEXED TO DNA

THE single-stranded DNA (ssDNA) binding protein gp32 from bacteriophag e T4 is essential for T4 DNA replication, recombination and repair. In vivo gp32 binds ssDNA as the replication fork advances and stimulates repulsion processivity and accuracy by a factor of several hundred(1) . Gp32 binding affects nearly every major aspect of DNA metabolism. Am ong its important functions are: (1) configuring ssDNA templates for e fficient use by the replisome including DNA polymerase; (2) melting ou t adventitious secondary structures; (3) protecting exposed ssDNA from nucleases; and (4) facilitating homologous recombination by binding s sDNA during strand displacement. We have determined the crystal struct ure of the gp32 DNA binding domain complexed to ssDNA at 2.2 Angstrom resolution. The ssDNA binding cleft comprises regions from three struc tural subdomains and includes a positively charged surface that runs p arallel to a series of hydrophobic pockets formed bg clusters of aroma tic side chains. Although only weak electron density is seen for the s sDNA, it indicates that the phosphate backbone contacts an electroposi tive cleft of the protein, placing the bases in contact with the hydro phobic pockets. The DNA mobility implied by the weak electron density may reflect the role of gp32 as a sequence-independent ssDNA chaperone allowing the largely unstructured ssDNA to slide freely through the c left.