DNA Replication (unzipping, unwounding, checking & replicating)
In molecular biology, DNA replication is the biological process of producing two identical replicas of DNA from one original DNAmolecule. This process occurs in all living organisms and is the basis for biological inheritance. The cell possesses the distinctive property of division, which makes replication of DNA essential.
DNA is made up of a double helix of two complementary strands. During replication, these strands are separated. Each strand of the original DNA molecule then serves as a template for the production of its counterpart, a process referred to as semiconservative replication.
As a result of semi-conservative replication, the new helix will be composed of an original DNA strand as well as a newly synthesized strand.
Cellular proofreading and error-checking mechanisms ensure near perfect fidelity for DNA replication.
In a cell, DNA replication begins at specific locations, or origins of replication, in the genome.
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Unwinding of DNA at the origin and synthesis of new strands, accommodated by an enzyme known as helicase, results in replication forks growing bi-directionally from the origin. A number of proteins are associated with the replication fork to help in the initiation and continuation of DNA synthesis. Most prominently, DNA polymerase synthesizes the new strands by adding nucleotides that complement each (template) strand. DNA replication occurs during the S-stage of interphase.
DNA replication (DNA amplification) can also be performed in vitro (artificially, outside a cell). DNA polymerases isolated from cells and artificial DNA primers can be used to initiate DNA synthesis at known sequences in a template DNA molecule. Polymerase chain reaction (PCR), ligase chain reaction (LCR), and transcription-mediated amplification (TMA) are examples.
DNA exists as a double-stranded structure, with both strands coiled together to form the characteristic double-helix. Each single strand of DNA is a chain of four types of nucleotides. Nucleotides in DNA contain a deoxyribose sugar, a phosphate, and a nucleobase. The four types of nucleotide correspond to the four nucleobasesadenine, cytosine, guanine, and thymine, commonly abbreviated as A, C, G and T. Adenine and guanine are purine bases, while cytosine and thymine are pyrimidines. These nucleotides form phosphodiester bonds, creating the phosphate-deoxyribose backbone of the DNA double helix with the nucleobases pointing inward (i.e., toward the opposing strand). Nucleobases are matched between strands through hydrogen bonds to form base pairs. Adenine pairs with thymine (two hydrogen bonds), and guanine pairs with cytosine (three hydrogen bonds).
DNA strands have a directionality, and the different ends of a single strand are called the “3′ (three-prime) end” and the “5′ (five-prime) end”. By convention, if the base sequence of a single strand of DNA is given, the left end of the sequence is the 5′ end, while the right end of the sequence is the 3′ end. The strands of the double helix are anti-parallel with one being 5′ to 3′, and the opposite strand 3′ to 5′. These terms refer to the carbon atom in deoxyribose to which the next phosphate in the chain attaches. Directionality has consequences in DNA synthesis, because DNA polymerase can synthesize DNA in only one direction by adding nucleotides to the 3′ end of a DNA strand.
The pairing of complementary bases in DNA (through hydrogen bonding) means that the information contained within each strand is redundant. Phosphodiester (intra-strand) bonds are stronger than hydrogen (inter-strand) bonds. This allows the strands to be separated from one another. The nucleotides on a single strand can therefore be used to reconstruct nucleotides on a newly synthesized partner strand.
- ^ The energetics of this process may also help explain the directionality of synthesis—if DNA were synthesized in the 3′ to 5′ direction, the energy for the process would come from the 5′ end of the growing strand rather than from free nucleotides. The problem is that if the high energy triphosphates were on the growing strand and not on the free nucleotides, proof-reading by removing a mismatched terminal nucleotide would be problematic: Once a nucleotide is added, the triphosphate is lost and a single phosphate remains on the backbone between the new nucleotide and the rest of the strand. If the added nucleotide were mismatched, removal would result in a DNA strand terminated by a monophosphate at the end of the “growing strand” rather than a high energy triphosphate. So strand would be stuck and wouldn’t be able to grow anymore. In actuality, the high energy triphosphates hydrolyzed at each step originate from the free nucleotides, not the polymerized strand, so this issue does not exist.
- ^ Pray, Leslie A. “Semi-Conservative DNA Replication; Meselson and Stahl”.
- ^ Imperfect DNA replication results in mutations. Berg JM, Tymoczko JL, Stryer L, Clarke ND (2002). “Chapter 27: DNA Replication, Recombination, and Repair“. Biochemistry. W.H. Freeman and Company. ISBN 0-7167-3051-0. External link in |chapter= (help)
- ^ Jump up to: a b Alberts B, Johnson A, Lewis J, Raff M, Roberts K, Walter P (2002). “5DNA Replication, Repair, and Recombination“. Molecular Biology of the Cell. Garland Science. ISBN 0-8153-3218-1. External link in |chapter= (help)
- ^ Jump up to: a b Berg JM, Tymoczko JL, Stryer L, Clarke ND (2002). “Chapter 27, Section 4: DNA Replication of Both Strands Proceeds Rapidly from Specific Start Sites“. Biochemistry. W.H. Freeman and Company. ISBN 0-7167-3051-0. External link in |chapter= (help)
- ^ Alberts, B., et al., Molecular Biology of the Cell, Garland Science, 4th ed., 2002, pp. 238–240 ISBN 0-8153-3218-1