DNA Topoisomerase
Functions of DNA Topoisomerases
There are a number of different types of topoisomerases, each specialising in a different aspect of DNA manipulation.
Accessing DNA
During transcription and DNA replication, the DNA needs to be unwound in order for the transcription/replication machinery to gain access to the DNA so it can be copied or replicate, respectively. Topoisomerase I can make single-stranded breaks to allow these processes to proceed.
Removing DNA Supercoils
During transcription and DNA replication, the DNA helix can become either over-wound or under-wound. For instance, during DNA replication, the progress of the replication fork generates positive supercoils ahead of the replication machinery and negative supercoils behind it. Such tensional problems also exist when transcribing DNA to make an RNA copy for protein synthesis. During these processes, the DNA can be supercoiled to such an extent that if left unchecked it could impede the progress of the protein machinery involved. This is prevented by topoisomerase I, which makes single-stranded nicks to relax the helix.
Strand Breakage during Recombination
Before the chromosomes separate from one another during cell division, they are able to exchange genetic information through a process known as recombination, where physical pieces of DNA on one chromosome can be swapped for DNA on the matching sister chromosome in order to shuffle the genetic information. Topoisomerase III can introduce single-strand breaks that are required for DNA to be exchanged by adjacent chromosomes.
Chromosome Condensation
During the cell cycle, chromosomes must be alternatively condensed and decondensed at specific stages. Topoisomerase II (gyrase) acts as a molecular motor, using the energy gained from ATP hydrolysis to introduce tight supercoils into the DNA helix in order to condense the chromosome. Because this process must be highly regulated, topoisomerase II can form molecular complexes with important cell cycle regulators (such as p53, TopBP1, 14-3-3 epsilon, and Cdc2) to ensure that chromosome condensation occurs at the correct time in the cell cycle.
Disentangling Intertwined DNA
During cell division, once the chromosomes have been replicated, they must separate and travel to opposite ends of the cell to become part of two separate daughter cells. Topoisomerases IV acts to disentangle the replicated daughter strands by making double-strand breaks that allow one duplex to pass through the other.
Topoisomerases as Drug Targets
Topoisomerases have been the focus for the treatment of certain diseases. Bacterial gyrase (topoisomerase II) and topoisomerase IV are the targets of two classes of antibiotic drugs: quinolones and coumarins. These antibiotics are used to treat an assortment of different diseases, such as pneumonia, tuberculosis and malaria, by inhibiting DNA replication in the bacteria responsible.
Eukaryotic topoisomerases I and II are the targets of an increasing number of anti-cancer drugs that act to inhibit these enzymes by blocking the reaction that reseals the breaks in the DNA. Often the binding of the drug is reversible, but if a replication fork runs into the blocked topoisomerase, then a piece of the gapped DNA strand not bound by the topoisomerase could be released, creating a permanent breakage in the DNA that leads to cell death. Most of these inhibitors are selective against either topoisomerase I or II, but some can target both enzymes.
Topoisomerase I inhibitors induce single-strand breaks into DNA, and can work by a variety of mechanisms. Some drugs, such as camptothecins, inhibit the dissociation of topoisomerase and DNA, leading to replication-mediated DNA damage, which can be repaired more efficiently in normal cells than in cancer cells (deficient for DNA repair). Topoisomerase I inhibitors can also cause gene inactivation through chromatid aberrations.
Topoisomerase II inhibitors, such as anthracyclines, are amongst the most widely used anti-cancer agents. These drugs are potent inducers of double strand breaks in DNA, and can cause arrest in the cell cycle at the G2 stage, the latter occurring by disrupting the interaction between topoisomerase II and regulators of the cell cycle, such as Cdc2. Topoisomerase II inhibitors can cause a wide range of chromosomal aberrations, and can act by either stabilising topoisomerase II-DNA complexes that are easily cleaved, or by interfering with the catalytic activity of the enzyme, both resulting in double-strand breaks in the DNA.
There are also dual inhibitors that target both topoisomerase I and II, which increases the potency of the anti-cancer effect. These drugs work by a variety of means: by recognising structural motifs present on both enzymes, by linking separate topoisomerase inhibitors together into a hybrid drug, or by using inhibitors that bind to and intercalate DNA.