These two strands serve as the template for the leading and lagging strands, which will be created as DNA polymerase matches complementary nucleotides to the templates; the templates may be properly referred to as the leading strand template and the lagging strand templates Leading strand The leading strand is the template strand of the DNA double helix so that the replication fork moves along it in the 3′ to 5′ direction. This allows the newly synthesized strand complementary to the original strand to be synthesized 5′ to 3′ in the same direction as the movement of the replication fork.

On the leading strand, a polymerase “reads” the DNA and adds nucleotides to it continuously. This polymerase is DNA polymerase III (DNA Pol III) in prokaryotes and presumably Pol ? [7][15] in yeasts. In human cells the leading and lagging strands are synthesized by Pol ? and Pol ? within the nucleus and Pol ? in the mitochondria. Pol ? can substitute for Pol ? in special circumstances. [16] Lagging strand The lagging strand is the strand of the template DNA double helix that is oriented so that the replication fork moves along it in a 5′ to 3′ manner.

Because of its orientation, opposite to the working orientation of DNA polymerase III, which moves on a template in a 3′ to 5′ manner, replication of the lagging strand is more complicated than that of the leading strand. On the lagging strand, primase “reads” the DNA and adds RNA to it in short, separated segments. In eukaryotes, primase is intrinsic to Pol ?. [17] DNA polymerase III or Pol ? lengthens the primed segments, forming Okazaki fragments. Primer removal in eukaryotes is also performed by Pol ?. 18] In prokaryotes, DNA polymerase I “reads” the fragments, removes the RNA using its flap endonuclease domain (RNA primers are removed by 5′-3′ exonuclease activity of polymerase I [weaver, 2005]), and replaces the RNA nucleotides with DNA nucleotides (this is necessary because RNA and DNA use slightly different kinds of nucleotides). DNA ligase joins the fragments together. Dynamics at the replication fork The assembled human DNA clamp, a trimer of the protein PCNA. As helicase unwinds DNA at the replication fork, the DNA ahead is forced to rotate.

This process results in a build-up of twists in the DNA ahead. [19] This build-up would form a resistance that would eventually halt the progress of the replication fork. DNA Gyrase is an enzyme that temporarily breaks the strands of DNA, relieving the tension caused by unwinding the two strands of the DNA helix; DNA Gyrase achieves this by adding negative supercoils to the DNA helix. [20] Bare single-stranded DNA tends to fold back on itself and form secondary structures; these structures can interfere with the movement of DNA polymerase.

To prevent this, single-strand binding proteins bind to the DNA until a second strand is synthesized, preventing secondary structure formation. [21] Clamp proteins form a sliding clamp around DNA, helping the DNA polymerase maintain contact with its template, thereby assisting with processivity. The inner face of the clamp enables DNA to be threaded through it. Once the polymerase reaches the end of the template or detects double-stranded DNA, the sliding clamp undergoes a conformational change that releases the DNA polymerase. Clamp-loading proteins are used to initially load the clamp, recognizing the junction between template and RNA