User:Nathan Harris/Tus: Difference between revisions

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The ''Ter'' region in ''E.coli'' between bases T5 and A9 is significantly underwound upon binding with Tus. This region of DNA is altered from standard B form which is attributed to straddling of ''Ter'' by interdomain β strands (βF and βG) and the L4 connecting loop of Tus. Tus interacts with ''Ter'' in a previously undescribed manner with β strands of Tus inserting almost perpendicularly into the major groove to recognise ''Ter''. Alteration of ''Ter'' is characterised by an extended major groove and a broadened minor groove generating an overall DNA bend of 20 degrees. Overall, contacts in these regions account for increased stability of the altered DNA shape and allow recognition of the appropriate ''Ter'' site <ref name = "Neylon" /><ref name = "Kamada" />.

=='''Mechanism of action'''==

The ability of Tus to terminate replication in ''E. coli'' in a polar manner is believed to involve the inhibition of DnaB helicase. This is achieved either through a “locked complex” model provided by Tus-Ter interactions providing a physical block, protein-protein interactions between Tus and DnaB, or through a combination of these two effects<ref name = "Kamada" /><ref name = "Kaplan" />.

The ability of Tus to terminate replication in ''E. coli'' in a polar manner is believed to involve the inhibition of DnaB helicase. This is achieved either through a “locked complex” model provided by Tus-Ter interactions providing a physical block, protein-protein interactions between Tus and DnaB, or through a combination of these two effects <ref name = "Kamada" /><ref name = "Kaplan" />.

===='''The Tus- ''Ter'' locked complex'''====

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==='''Tus-DnaB interactions'''===

Numerous studies support a model for replication termination resulting specifically from Tus-DnaB protein interactions.

Numerous studies support a model for replication termination resulting specifically from Tus-DnaB protein interactions. Experimentation in the field has demonstrated that the <scene name='User:Nathan_Harris/Tus/E49/1'>E49</scene> within the L1 loop of the non-permissive face of Tus is important in the formation of protein-protein interactions with DnaB. When this glutamic acid is exchanged for lysine (E49K), an increase in affinity for ''Ter'' and a decrease in affinity for DnaB result <ref name = "Henderson"> Henderson, T., Niles, A., Valjavec-Gratian, M. and Hill, T. (2001) Site-directed mutagenesis and phylogenetic comparisons of Escherichia coli Tus protein: DNA-protein interactions alone cannot account for Tus activity. Molecular Genetics and Genomics, 265 (6): 941-953.</ref><ref name = "Mulugu"> Mulugu, S., Potnis, A., Shamsuzzaman, T. J., Alexander, K. and Bastia, D. (2001) Mechanism of termination of DNA replication of Escherichia coli involves helicase-contrahelicase interaction. Proceedings of the National Academy of Science, USA, 98 (17): 9569-9574.</ref>. Despite the increased affinity for ''Ter'', this E49K mutatation results in a reduced capability of polar replication fork termination demonstrating the importance of Tus-DnaB interactions.

Experimentation in the field has demonstrated that the <scene name='User:Nathan_Harris/Tus/E49/1'>E49</scene> within the L1 loop of the non-permissive face of Tus is important in the formation of protein-protein interactions with DnaB. When this glutamic acid is exchanged for lysine (E49K), an increase in affinity for ''Ter'' and a decrease in affinity for DnaB result. Despite the increased affinity for ''Ter'', this E49K mutatation results in a reduced capability of polar replication fork termination demonstrating the importance of Tus-DnaB interactions.

In further confirmation of this helicase specific mechanism, the engineering of intra-strand covalent crosslinks introduced immediately upstream of the C6 of ''Ter'' prevent DnaB helicase from unwinding the C6. Despite this inability to unwind and from a locked complex with ''Tus'', polar fork termination is still permitted indicating that the formation of a locked complex is unnecessary for replication termination.

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Bastia, D., Zzaman, S., Krings, G., Saxena, M., Peng, X. and Greenberg, M. (2008) Replication termination mechanism as revealed by Tus-mediated polar arrest of a sliding helicase. Proceedings of the National Academy of Science, USA, 105 (93): 12831-12836.

Henderson, T., Niles, A., Valjavec-Gratian, M. and Hill, T. (2001) Site-directed mutagenesis and phylogenetic comparisons of Escherichia coli Tus protein: DNA-protein interactions alone cannot account for Tus activity. Molecular Genetics and Genomics, 265 (6): 941-953.

Mulugu, S., Potnis, A., Shamsuzzaman, T. J., Alexander, K. and Bastia, D. (2001) Mechanism of termination of DNA replication of Escherichia coli involves helicase-contrahelicase interaction. Proceedings of the National Academy of Science, USA, 98 (17): 9569-9574.

Neylon, C., Kralicek, A. V., Hill, T.M. and Dixon, N.E. (2005) Replication Termination in Escherichia coli: Structure and Antihelicase Activity of the Tus-Ter Complex. Microbiology and Molecular Biology, 69 (3): 501-526.

@@ Line 20: / Line 20: @@
 The ''Ter'' region in ''E.coli'' between bases T5 and A9 is significantly underwound upon binding with Tus. This region of DNA is altered from standard B form which is attributed to straddling of ''Ter'' by interdomain β strands (βF and βG) and the L4 connecting loop of Tus. Tus interacts with ''Ter'' in a previously undescribed manner with β strands of Tus inserting almost perpendicularly into the major groove to recognise ''Ter''. Alteration of ''Ter'' is characterised by an extended major groove and a broadened minor groove generating an overall DNA bend of 20 degrees. Overall, contacts in these regions account for increased stability of the altered DNA shape and allow recognition of the appropriate ''Ter'' site <ref name = "Neylon" /><ref name = "Kamada" />.
 =='''Mechanism of action'''==
-The ability of Tus to terminate replication in ''E. coli'' in a polar manner is believed to involve the inhibition of DnaB helicase.  This is achieved either through a “locked complex” model provided by Tus-Ter interactions providing a physical block, protein-protein interactions between Tus and DnaB, or through a combination of these two effects<ref name = "Kamada" /><ref name = "Kaplan" />.
+The ability of Tus to terminate replication in ''E. coli'' in a polar manner is believed to involve the inhibition of DnaB helicase.  This is achieved either through a “locked complex” model provided by Tus-Ter interactions providing a physical block, protein-protein interactions between Tus and DnaB, or through a combination of these two effects <ref name = "Kamada" /><ref name = "Kaplan" />.
 ===='''The Tus- ''Ter'' locked complex'''====
@@ Line 40: / Line 40: @@
 ==='''Tus-DnaB interactions'''===
-Numerous studies support a model for replication termination resulting specifically from Tus-DnaB protein interactions.
+Numerous studies support a model for replication termination resulting specifically from Tus-DnaB protein interactions. Experimentation in the field has demonstrated that the <scene name='User:Nathan_Harris/Tus/E49/1'>E49</scene> within the L1 loop of the non-permissive face of Tus is important in the formation of protein-protein interactions with DnaB. When this glutamic acid is exchanged for lysine (E49K), an increase in affinity for ''Ter'' and a decrease in affinity for DnaB result <ref name = "Henderson"> Henderson, T., Niles, A., Valjavec-Gratian, M. and Hill, T.  (2001) Site-directed mutagenesis and phylogenetic comparisons of Escherichia coli Tus protein: DNA-protein interactions alone cannot account for Tus activity.  Molecular Genetics and Genomics, 265 (6): 941-953.</ref><ref name = "Mulugu"> Mulugu, S., Potnis, A., Shamsuzzaman, T. J., Alexander, K. and Bastia, D.  (2001) Mechanism of termination of DNA replication of Escherichia coli involves helicase-contrahelicase interaction.  Proceedings of the National Academy of Science, USA, 98 (17): 9569-9574.</ref>. Despite the increased affinity for ''Ter'', this E49K mutatation results in a reduced capability of polar replication fork termination demonstrating the importance of Tus-DnaB interactions.
-Experimentation in the field has demonstrated that the <scene name='User:Nathan_Harris/Tus/E49/1'>E49</scene> within the L1 loop of the non-permissive face of Tus is important in the formation of protein-protein interactions with DnaB. When this glutamic acid is exchanged for lysine (E49K), an increase in affinity for ''Ter'' and a decrease in affinity for DnaB result. Despite the increased affinity for ''Ter'', this E49K mutatation results in a reduced capability of polar replication fork termination demonstrating the importance of Tus-DnaB interactions.
 In further confirmation of this helicase specific mechanism, the engineering of intra-strand covalent crosslinks introduced immediately upstream of the C6 of ''Ter'' prevent DnaB helicase from unwinding the C6. Despite this inability to unwind and from a locked complex with ''Tus'', polar fork termination is still permitted indicating that the formation of a locked complex is unnecessary for replication termination.
@@ Line 55: / Line 54: @@
 Bastia, D., Zzaman, S., Krings, G., Saxena, M., Peng, X. and Greenberg, M.  (2008) Replication termination mechanism as revealed by Tus-mediated polar arrest of a sliding helicase.  Proceedings of the National Academy of Science, USA, 105 (93): 12831-12836.
-Henderson, T., Niles, A., Valjavec-Gratian, M. and Hill, T.  (2001) Site-directed mutagenesis and phylogenetic comparisons of Escherichia coli Tus protein: DNA-protein interactions alone cannot account for Tus activity.  Molecular Genetics and Genomics, 265 (6): 941-953.
-Mulugu, S., Potnis, A., Shamsuzzaman, T. J., Alexander, K. and Bastia, D.  (2001) Mechanism of termination of DNA replication of Escherichia coli involves helicase-contrahelicase interaction.  Proceedings of the National Academy of Science, USA, 98 (17): 9569-9574.
 Neylon, C., Kralicek, A. V., Hill, T.M. and Dixon, N.E.  (2005) Replication Termination in Escherichia coli: Structure and Antihelicase Activity of the Tus-Ter Complex.  Microbiology and Molecular Biology, 69 (3): 501-526.