DNA polymerase



DNA polymerase is an enzyme which catalyzes the polymerization of Deoxyribonucleic acid (DNA). DNA polymerase is used in two ways by cells. First, it is used to synthesize a new DNA strand from an old DNA strand (or template strand). Before a cell divides into two cells, it must reproduce the genetic information for the new cell. Second, it is used to repair DNA. The genetic code can become corrupted during replication or by external mutagens leading to disfunctional proteins and even cell death. DNA polymerase contains a proofreading and repair DNA mechanism that is used to find and fix the errors.

Function
DNA polymerase is used in the DNA replication, the DNA repair, the DNA sequencing, and the Polymerase Chain Reaction. The function of DNA polymerase is adding it to synthesize with the end of 3' to form a new strand of DNA. To make the new strand, DNA polymerase needs primer which is a short single strand segment of DNA. The primer is known as a “starter” strand of DNA or RNA, because it is a starting material for synthesis to form the new DNA strand. During the replication, the errors could be made. DNA polymerase corrects the errors by synthesizing a new DNA strand. 

DNA Synthesis
DNA synthesis (or [DNA replication]) is copying DNA strands. In other words, it copies genetic information from DNA. This process is important. When DNA makes a new DNA, it uses DNA synthesis which copies the genetic materials from the template DNA strands. Every DNA is produced in this way.

Leading strands and lagging strands are used in the DNA synthesis. The leading strands are known as the DNA strands. They are pointing to “right” direction (5'->3') in a continuous manner. At the end of 3', DNA polymerase can be synthesized to build another strand because of the free OH group of 3'.

The lagging strands are located at the opposite sides of the leading strands. Obviously, they pointing to “left” direction (3'->5'). Unlikely, the DNA polymerase cannot build another strand in the “left” direction (3'->5'). Therefore, they synthesize with the Okazaki fragments which are making up the lagging strands in the replication. After this process, the primases are built. And then, the DNA polymerase can use the free OH group of 3' and synthesize to build another strand in 5'->3' direction. 

DNA Repairs

 * Main Article: DNA repair

Proofreading is one of DNA repair mechanisms of DNA polymerase. As DNA polymerase adds new nucleotides it can make errors which can induce mutations in the genetic code. Proofreading checks for errors made during replication.(Purves, p227) When proofreading finds out errors, it corrects them through one of the following mechanisms. However, sometimes errors are not recognized during proofreading, and can remain a permanent part of the genome.(Purves, p253)

Mismatch Repair
Mismatch repair is a mechanism which checks and fixes mismatched base pairs while replication, base-pairing could be mismatched. It is different from proofreading. Proofreading checks and corrects errors in the replication. In contrast, mismatch repair scans DNA and corrects mismatched base-pairing right after the replication.(Purves, p227)

Excision Repair
Excision repair is a mechanism which removes abnormal bases. Abnormal bases can be formed by chemical damage and replace with abnormal bases with normal (functional) bases. The mismatch repair is used for small portion of damage, but the excision repair is used for big portion of damage which cannot be repair by the mismatch repair. (Purves, p227)

Polymerase Chain Reaction
Polymerase Chain Reaction (PCR) is a laboratory technique that uses a heat stable DNA polymerase to make multiple copies of DNA. Today PCR is an essential technique in biotechnology. The technique involves the use repeated temperature cycling and increases the number of short region of DNA exponentially in a short time.

The cyclic process involves three steps. First, double-stranded helix structure of DNA is separated into one single strand of DNA by heat. A primer, a “starter” strand of DNA is added along with the heat stable DNA polymerase and four deoxyribonucleotide triphosphates such as dATP, dGTP, dCTP, and DTTP. Lastly, when cooled DNA polymerase copies the new strands of DNA. PCR requires base sequences at 3’ end of strand of DNA and two primers, about 15-20 base long, bind one region of DNA.

The heat-stable polymerase is required because to denature DNA, it should be heated to 90°C, but DNA polymerases are normally destroyed at this temperature. Thomas Brock first discovered a bacterium called Thermus aquaticus living in hot springs that can live at temperatures up to 95°C, and whose DNA polymerases do not denature. Kerry Mullis realized the usefullness of heat-resistant bacterium, and he added it during the cyclic process. For this work, he earned Nobel Prize and the technique greatly impacted biotechnology. The heat-stable DNA polymerase that was isolated from Thermus aquaticus and is now used in PCR is called Taq Polymerase.(Purves, p229-230)

DNA Sequencing
Besides PCR, Polymerase Chain Reaction, there is another important technique to determine the base of sequence of DNA. This technique is DNA sequencing. DNA sequencing is use of artificially altered nucleosides. Four letters, A, C, G, and T, are representing four nucleotide subunits of DNA strand. A is stand for adenine, C is stand for cytosine, G is stand for guanine, and T is stand for thymine. They are important subunits for the DNA sequencing, because they are carriers of genetic information. 

To do DNA sequencing, it should have four substances to work with it. Before use DNA sequencing, the fragments of DNA need to be denatured. After the fragments of DNA denatured, there will be only one single DNA strands. And then, the single strands will be put in the test tubes and mixed with four substances. Four substances are:
 * DNA polymerase which will help to synthesize the new strands,
 * Artificial primers which will start synthesize the new strands,
 * Four dNTP which are dATP, dGTP, dCTP, and dTTP, and
 * Four ddNTP (Purves, p231)

Prokaryotic DNA Polymerase
In the bacterium, five different kinds of the Prokaryotic DNA polymerase are found.
 * Pol I: Polymerase I was discovered by Arthur Kornberg in 1958, and it is known as DNA polymerase. It is useful polymerase for the DNA repair, because it has nick translation (5'->3') and proofreading (3'->5').
 * Pol II: Polymerase II is mostly used in the DNA repair. It is similar to polymerase I, because it has both nick translation (5'->3') and proofreading (3'->5').  In contrast to polymerase I, polymerase II is lack with the nick translation activity.
 * Pol III: Polymerase III is a main polymerase in the bacterium. It has with an ability of the proofreading (3'->5').
 * Pol IV: Polymerase IV is a Y family of DNA polymerase.
 * Pol V: Polymerase V is a Y family of DNA polymerase.

Eukaryotic DNA Polymerase
Eukaryotes have at least fifteen DNA polymerases. This five polymerases are the essential polymerases of the eukaryotic DNA polymerase.
 * Pol α: Polymerase α is a primase.
 * Pol β: Polymerase β is used in the DNA repair.
 * Pol γ: Polymerase γ replicates DNA.
 * Pol δ: Polymerase δ is a main polymerase in the eukaryotic DNA polymerases. It has an ability of the proofreading (3'->5').
 * Pol ε: Polymerase ε is a substitute polymerase of polymerase δ.

History
In 1959, Arthur Kornberg isolated the enzyme DNA polymerase from Escherichia coli. Later, in 1959, Kornberg was awarded the Nobel Prize in Physiology or Medicine for his work.

Related References

 * Purves, Sadava, Orians, and Heller. Life: The Science of Biology. Virginia: Gordonsville, 2004
 * DNA polymerase Wikipedia
 * DNA polymerase Answers.com
 * DNA replication Wikipedia
 * DNA sequencing Wikipedia

See Also:

 * Cell biology

DNA polimerase