Answer to Question #250802 in Microbiology for Pri

Question

DNA polymerase I (Pol I) of E. coli consists of three functional parts (domains): an N-terminal domain with 5´ to 3´ exonuclease activities required for removal of the RNA primer, a central domain responsible for 3´ to 5´ exonuclease proofreading, and a C-terminal domain with polymerase activity. Pol I is thought to simultaneously remove RNA primers and fill in the gaps that result. A group of proteins known as RNaseH also have 5´ to 3´ exonuclease activity and can thus remove RNA primers. However, they lack the other two functions observed for Pol I. Predict the ability of the following mutants to replicate DNA:

(1) a strain with a mutant gene encoding Pol I such that it no longer has polymerase activity (but retains both types of nuclease activities);   

(2) a strain without RNaseH proteins;

Answer 1

1) DNA polymerase 3 is required for leading and lagging strand replication, whereas DNA polymerase 1 is required for removing RNA primers from fragments and replacing them with the required nucleotides. These enzymes cannot be substituted for one another because they perform different functions.

2) During DNA replication, repair, and recombination, DNA polymerase I fills gaps in the DNA. Editing and proofreading are also performed by DNA polymerase II, mostly on the lagging strand. The major replicative enzyme is DNA polymerase III.