BINDING-SPECIFICITY OF T4 DNA-POLYMERASE TO RNA

Bacteriophage T4 DNA polymerase, product of phage gene 43 (gp43), is a multifunctional DNA-binding protein and a key component of the phage DNA replicase. It is also an RNA-binding protein that selectively reco gnizes a site on its mRNA (the translational operator) and represses i ts own translation. We examined the ability of the protein to discrimi nate between DNA and RNA by using a gel mobility shift assay with defi ned RNA and DNA substrates. A higher affinity to RNA as compared with DNA (about 100-fold) was observed in assays that utilized synthetic DN A and in vitro transcribed RNA substrates bearing the T4 gene 43 trans lational operator sequence. The replacement of thymine with uracil in the synthetic DNA did not improve binding. The results suggest that th e protein's selectivity for RNA is based in structure (intramolecular interactions) specific to the ribonucleotide sequence of the operator. Competition studies suggest that the protein determinants for RNA and DNA recognition are only partially overlapping.