Structure/Function Relationships in the Bacteriophage T4 Single-Stranded DNA-Binding Protein

Abstract

Bacteriophage T4 gene 32 protein (gp32) is a prototype for a class of proteins whose fundamental feature includes a very strong affinity for single-stranded nucleic acids. This binding appears to be largely nonspecific with respect to sequence and favors single-stranded over doublestranded nucleic acids. Tight binding to single-stranded nucleic acids is undoubtedly important for the essential role of gp32 in T4 DNA replicaition, repair, and recombination [for a review see 1]. When E. coli, infected with T4 bacteriophage carrying temperature-sensitive mutations in gene 32, is shifted to a nonpermissive temperature, T4 DNA replication ceases [2], and the intracellular T4 DNA is rapidly degraded to small fragments [3]. The unbound concentration of gp32 in vivo is in the range of 2 to 3 µM [4], and it is thus present in “stoichiometric” amounts relative to the number of replication forks present. This finding is consistent with the presumed role of gp32 in T4 DNA replication which is to remove adventitious secondary structures from the single-stranded DNA template just ahead of the advancing replication fork and to protect the resulting ssDNA from nuclease degradation. In vitro DNA replication assays carried out with T4 DNA polymerase (gp43) and three T4 DNA polymerase accessory proteins (gp44, gp45, and gp62) indicate that the addition of gp32 stimulates the rate of DNA synthesis by 100- to 200-fold [5,6].