PROCESSIVITY OF THE GENE-41 DNA HELICASE AT THE BACTERIOPHAGE-T4 DNA-REPLICATION FORK

The gene 41 protein is the DNA helicase associated with the bacterioph age T4 DNA replication fork. This protein is a major component of the primosome, being essential for coordinated leading and lagging strand DNA synthesis, Models suggest that such DNA helicases are loaded only onto DNA at origins of replication, and that they remain with the ensu ing replication fork until replication is terminated. To test this ide a, we have measured the extent of processivity of the 41 protein in th e context of an in vitro DNA replication system composed of eight puri fied proteins (the gene 43, 44/62, 45, 32, 41, 59, and 61 proteins), A fter starting DNA replication in the presence of these proteins, we di luted the 41 helicase enough to prevent any association of new helicas e molecules and analyzed the replication products, We measured an asso ciation half-life of 11 min, revealing that the 41 protein is processi ve enough to finish replicating the entire 169-kilobase T4 genome at t he observed replication rate of similar to 400 nucleotides/s. This pro cessivity of the 41 protein does not require the 59 protein, the prote in that catalyzes 41 protein assembly onto 32 protein-covered single-s tranded DNA, The stability we measure for the 41 protein as part of th e replication fork is greater than estimated for it alone on single-st randed DNA, We suggest that the 41 protein interacts with the polymera se holoenzyme at the fork, both stabilizing the other protein componen ts and being stabilized thereby.