Summary: Transposase Tn5 dimerisation domain
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Transposase Edit Wikipedia article
Transposase is an enzyme that binds to the ends of a transposon and catalyzes the movement of the transposon to another part of the genome by a cut and paste mechanism or a replicative transposition mechanism.
The word "transposase" was first coined by the individuals who cloned the enzyme required for tranposition of the Tn3 transposon. The existence of transposons was postulated in the late 1940s by Barbara McClintock, who was studying the inheritance of maize, but the actual molecular basis for transposition was described by later groups. McClintock discovered that pieces of the chromosomes changed their position, jumping from one chromosome to another. The repositioning of these transposons (which coded for color) allowed other genes for pigment to be expressed. Transposition in maize causes changes in color; however, in other organisms, such as bacteria, it can cause antibiotic resistance. Transposition is also important in creating genetic diversity within species and adaptability to changing living conditions. During the course of human evolution, as much as 40% of the human genome has moved around via methods such as transposition of transposons.
Transposases are classified under EC number EC 2.7.7.
Genes encoding transposases are widespread in the genomes of most organisms and are the most abundant genes known.
 Transposase Tn5
|Transposase Tn5 dimerisation domain|
|tn5 transposase: 20mer outside end 2 mn complex|
Transposase Tn5 is a member of the RNase superfamily of proteins which includes retroviral integrases. Tn5 can be found in Shewanella and Escherichia bacteria. The transposon codes for antibiotic resistance to kanamycin and other aminoglycoside antibiotics.
Tn5 and other transposases are notably inactive. Because DNA transposition events are inherently mutagenic, the low activity of transposases is necessary to reduce the risk of causing a fatal mutation in the host, and thus eliminating the transposable element. One of the reasons Tn5 is so unreactive is because the N- and C-termini are located in relatively close proximity to one another and tend to inhibit each other. This was elucidated by the characterization of several mutations which resulted in hyperactive forms of transposases. One such mutation, L372P, is a mutation of amino acid 372 in the Tn5 transposase. This amino acid is generally a leucine residue in the middle of an alpha helix. When this leucine is replaced with a proline residue the alpha helix is broken, introducing a conformational change to the C-Terminal domain, separating it from the N-Terminal domain enough to promote higher activity of the protein. The transposition of a transposon often needs only three pieces: the transposon, the transposase enzyme, and the target DNA for the insertion of the transposon. This is the case with Tn5, which uses a cut-and-paste mechanism for moving around transposons.
Tn5 and most other transposases contain a DDE motif, which is the active site that catalyzes the movement of the transposon. Aspartate-97, Aspartate-188, and Glutamate-326 make up the active site, which is a triad of acidic residues. The DDE motif is said to coordinate divalent metal ions, most often magnesium and manganese, which are important in the catalytic reaction. Because transposase is incredibly inactive, the DDE region is mutated so that the transposase becomes hyperactive and catalyzes the movement of the transposon. The glutamate is transformed into an aspartate and the two asparates into glutamates. Through this mutation, the study of Tn5 becomes possible, but some steps in the catalytic process are lost as a result.
There are several steps which catalyze the movement of the transposon, including Tnp binding, synapsis (the creation of a synaptic complex), cleavage, target capture, and strand transfer. Transposase then binds to the DNA strand and creates a clamp over the transposon end of the DNA and inserts into the active site. Once the transposase binds to the transposon, it produces a synaptic complex in which two transposases are bound in a cis/trans relationship with the transposon.
In cleavage, the magnesium ions activate oxygen from water molecules and expose them to nucleophilic attack. This allows the water molecules to nick the 3' strands on both ends and create a hairpin formation, which separates the transposon from the donor DNA. Next, the transposase moves the transposon to a suitable location. Not much is known about the target capture, although there is a sequence bias which has not yet been determined. After target capture, the transposase attacks the target DNA nine base pairs apart, resulting in the integration of the transposon into the target DNA.
As mentioned before, due to the mutations of the DDE, some steps of the process are lost—for example, when this experiment is performed in vitro, and SDS heat treatment denatures the transposase. However, it is still uncertain what happens to the transposase in vivo.
The study of transposase Tn5 is of general importance because of its similarities to HIV-1 and other retroviral diseases. By studying Tn5, much can also be discovered about other transposases and their activities.
 Sleeping Beauty transposase
The Sleeping Beauty (SB) transposase is the recombinase that drives the Sleeping Beauty transposon system. SB transposase belongs to the DD[E/D] family of transposases, which in turn belong to a large superfamily of polynucleotidyl transferases that includes RNase H, RuvC Holliday resolvase, RAG proteins, and retroviral integrases. The SB system is used primarily in vertebrate animals for gene transfer, including gene therapy, and gene discovery The recently engineered SB100X is an especially powerful enzyme that directs the highest levels of transposon integration yet developed
- Cell. 1979 Dec;18(4):1153-63.
- Reznikoff, William S. (2003). "Tn5 as a model for understanding DNA transposition". Molecular Microbiology 47 (5): 1199–1206. doi:10.1046/j.1365-2958.2003.03382.x. PMID 12603728.
- Aziz, R.K., M. Breitbart and R.A. Edwards (2010). Transposases are the most abundant, most ubiquitous genes in nature. Nucleic Acid Research 38(13): 4207-4217.
- Lovell, Scott; Scott Lovell; Igor Y. Goryshin; William R. Reznikoff; Ivan Rayment (2002). "Two-metal active site binding of a Tn5 transposase synaptic complex". Nature Structural Biology 9 (4): 278–81. doi:10.1038/nsb778. PMID 11896402.
- Peterson, G; Gregory Peterson; William Reznikoff (2003). "Tn5 Transposase Active Site Mutations Suggest Positions of Donor Backbone DNA in Synaptic Complex". The Journal of Biological Chemistry 278 (3): 1904–1909. doi:10.1074/jbc.M208968200. PMID 12424243.
- Ivics, Z., P.B. Hackett, R.A. Plasterk, and Z. Izsvak (1997). Molecular reconstruction of Sleeping Beauty: a Tc1-like transposon from fish and its transposition in human cells. Cell 91: 501-510.
- Craig,N.L. (1995) Unity in transposition reactions. Science 270: 253-254.
- Nesmelova, I.V. and P.B. Hackett (2010) DDE Transposases: structural similarity and diversity. Adv. Drug Del. Rev. 62: 1187-1195.
- Ivics, Z., Izsvak, Z. (2005) A whole lotta jumpin’ goin’ on: new transposon tools for vertebrate functional genomics. Trends Genet. 21: 8-11.
- Izsvak, Z., Hackett, P.B., Cooper, L.J.N., and Ivics, Z. (2010). Translating Sleeping Beauty transposition to molecular therapy: victories and challenges. BioEssays, 32: 756-767.
- Aronovich, E.L., McIvor, R.S., and Hackett, P.B. (2011). The Sleeping Beauty transposon system – A non-viral vector for gene therapy. Hum. Mol. Genet. (in press)
- Carlson, C.M., Largaespada, D.A. (2005). Insertional mutagenesis in mice: new perspectives and tools. Nature Rev. Genet. 6: 568-580.
- Copeland, N.G., Jenkins, N.A. (2010). Harnessing transposons for cancer gene discovery. Nature Rev. Genet. 10: 696-706.
- Mátés, L., et al. (2009) Molecular evolution of a novel hyperactive Sleeping Beauty transposase enables robust stable gene transfer in vertebrates. Nature Genet. 41: 753-761
- Grabundzija, I., Irgang, M., Mátés, L., Belay, E., Matrai, J., Gogol-Döring, A., Kawakami, K., Chen, W., Ruiz, P., Chuah, M.K., VandenDriessche, T., Izsvák, Z., Ivics, Z. (2010). Comparative analysis of transposable element vector systems in human cells. Mol. Ther. 18: 1200-1209.
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Transposase Tn5 dimerisation domain Provide feedback
Transposons are mobile DNA sequences capable of replication and insertion into the chromosome. Typically transposons code for the transposase enzyme, which catalyses insertion, found between terminal inverted repeats. Tn5 has a unique method of self- regulation in which a truncated version of the transposase enzyme acts as an inhibitor . The catalytic domain of the Tn5 transposon is found in PF01609. This domain mediates dimerisation in the known structure.
Davies DR, Braam LM, Reznikoff WS, Rayment I; , J Biol Chem 1999;274:11904-11913.: The three-dimensional structure of a Tn5 transposase-related protein determined to 2.9-A resolution. PUBMED:10207011 EPMC:10207011
External database links
This tab holds annotation information from the InterPro database.
InterPro entry IPR003201
Transposons are mobile DNA sequences capable of replication and insertion into the chromosome. Typically transposons code for the transposase enzyme, which catalyses insertion, found between terminal inverted repeats [PUBMED:6306482]. Tn5 has a unique method of self- regulation in which a truncated version of the transposase enzyme acts as an inhibitor [PUBMED:10207011].
More information about these proteins can be found at Protein of the Month: Transposase [PUBMED:].
Below is a listing of the unique domain organisations or architectures in which this domain is found. More...
The graphic that is shown by default represents the longest sequence with a given architecture. Each row contains the following information:
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|Seed source:||Pfam-B_5683 (release 5.2)|
|Author:||Mian N, Bateman A|
|Number in seed:||2|
|Number in full:||403|
|Average length of the domain:||97.40 aa|
|Average identity of full alignment:||52 %|
|Average coverage of the sequence by the domain:||31.63 %|
|HMM build commands:||
build method: hmmbuild -o /dev/null HMM SEED
search method: hmmsearch -Z 23193494 -E 1000 --cpu 4 HMM pfamseq
|Family (HMM) version:||11|
|Download:||download the raw HMM for this family|
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The tree shows the occurrence of this domain across different species. More...
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For all of the domain matches in a full alignment, we count the number that are found on all sequences in the alignment. This total is shown in the purple box.
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There is 1 interaction for this family. More...
We determine these interactions using iPfam, which considers the interactions between residues in three-dimensional protein structures and maps those interactions back to Pfam families. You can find more information about the iPfam algorithm in the journal article that accompanies the website.
For those sequences which have a structure in the Protein DataBank, we use the mapping between UniProt, PDB and Pfam coordinate systems from the PDBe group, to allow us to map Pfam domains onto UniProt sequences and three-dimensional protein structures. The table below shows the structures on which the Dimer_Tnp_Tn5 domain has been found. There are 6 instances of this domain found in the PDB. Note that there may be multiple copies of the domain in a single PDB structure, since many structures contain multiple copies of the same protein seqence.
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