Replication Fork

The replication fork* is a region where a cell's DNA* double helix has been unwound and separated to create an area where DNA polymerases and the other enzymes involved can use each strand as a template to synthesize a new double helix.

An enzyme called a helicase* catalyzes strand separation. Once the strands are separated, a group of proteins called helper proteins prevent the strands from coming back together.

DNA polymerase can not create new polymers. The enzyme can only extend existing strands by adding new nucleotides* to the 3'-hydroxyl end of an existing polymer. So before DNA polymerase can begin working, primase* (a type of RNA polymerase) binds to each strand of DNA at the replication fork and synthesizes a short (3 to 10 base) strand of RNA. This short RNA polymer called a primer provides a strand end for DNA polymerase to add bases to.

Since DNA polymerases can only add nucleotides to the 3'-hydroxyl end of a nucleotide polymer, and the two strands of the original DNA helix are oriented in opposite directions - synthesis of new polymers has to proceed in opposite directions on each of the two template strands at the replication fork.

In one direction, DNA is replicated as one continuous strand. This is called the leading strand. The other strand is called the lagging strand.

On the lagging strand, the new strand's 3'-hydroxyl end points away from the replication fork. This forces the elongation process to occur in a discontinuous manner. As replication moves along the template strand, a series of shorter DNA polymers form. Each stretch is initiated with its own RNA primer.

The shorter lengths of double-stranded DNA formed along the lagging strand are called Okazaki fragments.

DNA polymerase III performs most of the synthesis activity, but when an Okazaki fragment extends to the point that it overlaps with the previous RNA primer, RNA nucleotides are removed and replaced by DNA. This requires DNA Polymerase I, which has exonuclease activity.

Once the RNA primer is completely replaced by DNA, the two DNA fragments are joined by a ligase enzyme.

*The process is quite complex and involves numerous enzymes and helper proteins. This discussion is limited to the major enzymes involved.

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