Sand box 211: Difference between revisions
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== '''Description''' == | == '''Description''' == | ||
[[Image:activity.jpg | thumb | left | The different substrates of the T5 5'-exonuclease]]The T5 5'-exonuclease, also called T5 5'-3' exonuclease, is a member of the family of flap endonucleases (FEN), also known as 5'-nucleases, and is composed of 291 amino acids. Flap endonucleases are present in almost all living organisms. They participate in DNA replication, by removing the Okazaki fragments, and repair processes. In addition, they are able to cleave branched DNA by catalyzing the exonucleolytic hydrolysis of the phosphodiester bonds present in the DNA. Furthermore, they have an endonucleolytic activity which consists in cleaving DNA flap structures. Both activities are structure-specific because they only take place in presence of double strand-single strand junctions in bifurcated nucleic acid substrates like the flap (A on the figure on the left), the pseudo-Y (B on the figure on the left) and the 5'-overhanging hairpin (C on the figure on the left) substrates. | [[Image:activity.jpg | thumb | left | The different substrates of the T5 5'-exonuclease]]The '''T5 5'-exonuclease''', also called '''T5 5'-3' exonuclease''', is a member of the family of flap endonucleases (FEN), also known as 5'-nucleases, and is composed of 291 amino acids. Flap endonucleases are present in almost all living organisms. They participate in DNA replication, by removing the Okazaki fragments, and repair processes. In addition, they are able to cleave branched DNA by catalyzing the exonucleolytic hydrolysis of the phosphodiester bonds present in the DNA. Furthermore, they have an endonucleolytic activity which consists in cleaving DNA flap structures. Both activities are structure-specific because they only take place in presence of double strand-single strand junctions in bifurcated nucleic acid substrates like the flap (A on the figure on the left), the pseudo-Y (B on the figure on the left) and the 5'-overhanging hairpin (C on the figure on the left) substrates. | ||
The T5 5'-exonuclease is found in bacteriophages T5 and is coded by the gene D15. It is called metalloenzyme because it has binding sites for divalent metal ions without which the enzyme isn't able to cut DNA <ref>PMID:12606565</ref>. However, it can bind to DNA without binding a divalent metal ion. | The T5 5'-exonuclease is found in bacteriophages T5 and is coded by the gene D15. It is called metalloenzyme because it has binding sites for divalent metal ions without which the enzyme isn't able to cut DNA <ref>PMID:12606565</ref>. However, it can bind to DNA without binding a divalent metal ion. | ||
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== '''Structure''' == | == '''Structure''' == | ||
<Structure load='1EXN' size='350' frame='true' align='right' caption='T5 phage exonuclease' scene='Insert optional scene name here' /> | <Structure load='1EXN' size='350' frame='true' align='right' caption='T5 phage exonuclease (PDB code [[1exn]])' scene='Insert optional scene name here' /> | ||
<scene name='Sand_box_211/1exn/4'>T5 5'-exonuclease</scene> is a homodimeric protein composed of two identical chains,<scene name='Sand_box_211/Vghjvjh/4'>chain a</scene> and <scene name='Sand_box_211/Vhj/3'>chain b</scene>. | <scene name='Sand_box_211/1exn/4'>T5 5'-exonuclease</scene> is a homodimeric protein composed of two identical chains,<scene name='Sand_box_211/Vghjvjh/4'>chain a</scene> and <scene name='Sand_box_211/Vhj/3'>chain b</scene>. | ||
Both chains contain a hole, bound by a <scene name='Celina_Pinto/Sandbox_211/Helical_arch/2'>helical arch</scene> composed of two helices in which <scene name='Celina_Pinto/Sandbox_211/Residues_helical_arch/2'>hydrophobic and positively charged residues</scene> are located. The helical arch is situated in front of the active site and only single-stranded DNA can pass through it<ref>PMID:8657312</ref>. Since the enzyme is able to cleave double-stranded DNA, the enzyme has a conformational flexibility to facilitate DNA threading which is required to process the 5' nuclease substrates in the active site. | Both chains contain a hole, bound by a <scene name='Celina_Pinto/Sandbox_211/Helical_arch/2'>helical arch</scene> composed of two helices in which <scene name='Celina_Pinto/Sandbox_211/Residues_helical_arch/2'>hydrophobic and positively charged residues</scene> are located. The helical arch is situated in front of the active site and only single-stranded DNA can pass through it<ref>PMID:8657312</ref>. Since the enzyme is able to cleave double-stranded DNA, the enzyme has a conformational flexibility to facilitate DNA threading which is required to process the 5' nuclease substrates in the active site. | ||