RTP and Tus: Difference between revisions
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The structure of Tus is unusual for a DNA-binding protein. It binds ''Ter'' DNA as an asymmetrical monomer, which establishes the basis for its polar arrest of the replication fork. Tus has three distinct regions: two α-helical regions and central β-strands which jointly form a large, positively-charged central cleft (Kamada, 1996). The core β-structures embrace 13 base pairs of duplex DNA by partial insertion into the major groove, and at least 30 other residues make nonspecific contacts with the DNA backbone. | The structure of Tus is unusual for a DNA-binding protein. It binds ''Ter'' DNA as an asymmetrical monomer, which establishes the basis for its polar arrest of the replication fork. Tus has three distinct regions: two α-helical regions and central β-strands which jointly form a large, positively-charged central cleft (Kamada, 1996). The core β-structures embrace 13 base pairs of duplex DNA by partial insertion into the major groove, and at least 30 other residues make nonspecific contacts with the DNA backbone. | ||
The positioning of α-helices in the Tus protein is particularly interesting. Two protrude from both the amino and carboxy domains to clasp the DNA duplex, thereby shielding the interdomain β structures from direct contacts with other proteins (such as the DnaB helicase). The concentration of α-helices on the non-permissive face of Tus is absolutely cruical to the protein's ability to form a locked complex with the ''Ter'' site. More about this later! | The positioning of α-helices in the Tus protein is particularly interesting. Two protrude from both the amino and carboxy domains to clasp the DNA duplex, thereby shielding the interdomain β structures from direct contacts with other proteins (such as the DnaB helicase). The concentration of α-helices on the non-permissive face of Tus is absolutely cruical to the protein's ability to form a locked complex with the ''Ter'' site. More about this later! | ||
'''So how does Tus actually stop the replication fork? And why is it a polar arrest mechanism?''' | '''So how does Tus actually stop the replication fork? And why is it a polar arrest mechanism?''' | ||
Revision as of 13:14, 13 May 2011
A comparison of the Replication Terminator Protein (from Bacillus subtillis) and Tus (from Escerishia coli) provides an interesting insight into how proteins with vastly different structures and mechanisms of action can produce essentially identical effects in their native systems.
Looking at the structures of these two proteins, it is not immediately obvious that they would perfom the same function, specifically, to arrest the progression of the replication fork along the bacterial chromosome at specific sites (termed Ter sites). Furthermore, this arrest-mechanism functions in a polar manner in both organisms, which is perhaps surprising considering the symmetrical characteristics of both proteins.
The structure of Tus is unusual for a DNA-binding protein. It binds Ter DNA as an asymmetrical monomer, which establishes the basis for its polar arrest of the replication fork. Tus has three distinct regions: two α-helical regions and central β-strands which jointly form a large, positively-charged central cleft (Kamada, 1996). The core β-structures embrace 13 base pairs of duplex DNA by partial insertion into the major groove, and at least 30 other residues make nonspecific contacts with the DNA backbone. The positioning of α-helices in the Tus protein is particularly interesting. Two protrude from both the amino and carboxy domains to clasp the DNA duplex, thereby shielding the interdomain β structures from direct contacts with other proteins (such as the DnaB helicase). The concentration of α-helices on the non-permissive face of Tus is absolutely cruical to the protein's ability to form a locked complex with the Ter site. More about this later!
So how does Tus actually stop the replication fork? And why is it a polar arrest mechanism?
Mulcair et al (2006) discovered that the key to Tus forming a locked complex with Ter was twofold: firstly, the locked complex was formed only on the approach of DnaB helicase (the leading edge of the replication fork), and secondly, this locked complex was due to the base-flipping of C6 of Ter DNA into a cytosine-specific binding pocket on Tus. The approach of DnaB is essential to lock formation as strand separation is required before the C6 base can twist out of the helix. This C6 binds somewhere near the α4 helix, in or near the DNA-binding channel.
His144 is a particularly important residue - it exists as its conjugate acid in the locked complex, forming hydrogen bonds with C6. Other residues - for example Phe140 and Gly149 - are also strictly conserved amongst different species' Tus protiens; many of the conserved residues among different Ter sites make base-specific contacts with Tus.
The locked Tus-Ter complex is the most stable known monomeric DNa binding protein with a double-stranded sequence-specific recognition sequence - a half life of 550min has been reported (Mulcair, 2006). The formation of a large hydrogen-bond network is critical to sequence recognition and the stability of the twisted β-strands lying across the major groove.
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