The replication of a DNA molecule involves polymerization of special energy-carrying nucleotides called triphosphate deoxyribonucleotides since they are bound to three phosphate groups. The energy released by the enzymatic removal of two of the phosphates is utilized in the linking of each nucleotide to its neighbor on the growing DNA nucleoside.
DNA replication begins at a specific area along the molecule called the origin of replication. At the origin, histones are removed to expose the DNA strand, then the enzyme helicase untwists the replicating portion of the molecule and breaks the hydrogen bonds between complementary base pairs, causing the formation of a replication fork in the direction of replication. Stabilizing proteins keep the separated strands from twing back up.
An enzyme called RNA polymerase (primase) begins the replication process by adding RNA nucleotides to each template nucleoside. A molecule of DNA polymerase III binds to each of the separated strands. This enzyme adds nucleotide bases to their complementary bases on the template strand after the RNA primer sequence and proofreads to prevent improper nucleotides from being joined tothe template. DNA polymerase is directional so it will only completely build and proofread the nucleoside that moves in the 5' to 3' direction. This is called the continuous (leading) strand. Following DNA polymerase III, a new enzyme called DNA polymerase I removes the RNA primers and replaces them with DNA nucleotides.
The nucleoside that moves from 3' to 5' is called the discontinuous (lagging) strand since DNA polymerase III cannot continuously add nucleotides in that direction. Instead, primase adds RNA bases in several places along the growing strand, enabling DNA polymerase III to add DNA nucleotides between them. These completed DNA portions are called Okazaki fragments. DNA polymerase I replaces the RNA with DNA along the chain, filing the gaps between Okazaki fragments, but leaves unconnected "nicks" (unjoined regions) in the sugar-phosphate backbone. To complete the strand, a new molecule called DNA ligase links the nicks together. DNA gyrase twists the new double helix back into a supercoiled form in the bacterial cell.
Bacterial DNA replication is bidirectional since the chromosome is circular. It begins from a central origin and proceeds around the chromosome until the two polymerase enzymes meet. The torsion placed on the separated strands by the untwisting activity of helicase is relaxed by the enzyme topoisomerase by cutting the twisting sections and re-joining them opposite to the direction of the supercoil.
Another aspect of the replication of bacterial DNA is the process of methylation. During replication, some of the nucleotides (usually adenine, seldom cytosine) along specific sequences on the new strand have methyl groups (-CH3) added. This process is necessary for several reasons:
1. Methylated sequences serve to block or turn
on the transcription of mRNA along gene sequences,
thus act as a control mechanism for genetic expression.
2. These sequences may serve as initiators for DNA replication to begin.
3. Since viruses do not have methylated sequences
along their genome, the presence of these may
allow bacteria to seek out and enzymatically degrade bacteriophage prophage sequences.
4. Methylation may be involved in the process
of DNA repair.
Eukaryote DNA replication proceeds in a manner very similar to that of bacteria, with the following exceptions:
A. Eukaryotes utilize four different DNA polymerase
which initiates synthesis and
places primers (bacteria use primase for this), s which elongates the leading strand, e which
elongates the lagging strand and g that replicates mitochondrial DNA (note - mitchondrial
DNA is circular and naked in the mitochondrial matrix).
B. Eukaryote DNA requires many points of replication owing to its large size.
C. Eukaryote Okazaki fragments are far shorter than those of prokaryotes.
D. Methylation of plant and animal DNA occurs only on cytosine molecules.