DNA binding proteins can have a variety of motifs. The Catabolite Activator Protein (CAP) has a helix-turn-helix motif in it's DNA binding domain. The same protein is also known as Cyclic AMP Receptor Protein (CRP) and is coded for by the crp gene in E. coli. Cyclic AMP (cAMP - Magenta in the structure) binds to the protein, which then binds to DNA, activating transcription. The protein binds to a 22bp site, which consists of two groups of crucial bases, separated by one group of 10 bases (one turn of a DNA helix). Structure of CAP This protein has two domains: the N-terminal domain (red) which binds cAMP, and the C-terminal domain (yellow) which binds DNA. It is the latter domain which contains the helix-turn-helix motif. One helix is the recognition helix (red). It is stabilised by hydrophobic interactions with the other helix (yellow), which is almost perpendicular. The protein binds as a dimer, and dimerisation results in two recognition helices being 3.4nm apart (one turn of DNA). This is another common feature seen in many DNA binding proteins. Recognition by CAP The two-fold symmetry of the protein (after dimerisation ) once again matches the 2-fold symmetry of the target sequence. It must be noted that not ALL target sequences are symmetrical, but if they are, it is likely the protein which recognises the site will be also. Most proteins which recognise symetrical sites act as dimers and the dimer interaction leads to the symetry of the dimer. Each subunit of the dimer makes approximately 12-13 non-specific (ionic) interactions with the backbone of the DNA. The specific interactions are made by ARG 180, 185, (magenta) and GLU 181 (cyan) hydrogen bonding to the bases in the TGTGA sequence. So the protein is probably guided into place by non-specific interactions, and then "locked" in by specific ones. The cAMP (orange) bound to the CAP monomers changes the conformation of the dimer so that the distance between the recognition helices matches the spacing required for the binding. When cAMP is not bound the distance between the helices changes and the spacing is no longer optimal. The DNA is bent when the CAP protein binds, the overall bending being greater than 90o. This appears to be essential for the correct fit of the protein. It is also important in activating transcription; although it is not known whether the bending helps unwind the DNA, or position the RNA Polymerase correctly. One might speculate that the kinking of the DNA helps release the CAP protein from the DNA when the [cAMP} falls. Once the cAMP leaves the CAP the energy involved in the kinking may force the unliganded (no cAMP) protein off the DNA binding sequence. (A little bit of speculation by JWT) There are other motifs involved in DNA binding proteins, and these are dealt with in the last two tutorials on this issue. Press NEXT to view them now, or go HOME if you wish to finish for the day.
Cyclic AMP (cAMP - Magenta in the structure) binds to the protein, which then binds to DNA, activating transcription.
The protein binds to a 22bp site, which consists of two groups of crucial bases, separated by one group of 10 bases (one turn of a DNA helix).
Structure of CAP This protein has two domains: the N-terminal domain (red) which binds cAMP, and the C-terminal domain (yellow) which binds DNA. It is the latter domain which contains the helix-turn-helix motif. One helix is the recognition helix (red). It is stabilised by hydrophobic interactions with the other helix (yellow), which is almost perpendicular. The protein binds as a dimer, and dimerisation results in two recognition helices being 3.4nm apart (one turn of DNA). This is another common feature seen in many DNA binding proteins. Recognition by CAP The two-fold symmetry of the protein (after dimerisation ) once again matches the 2-fold symmetry of the target sequence. It must be noted that not ALL target sequences are symmetrical, but if they are, it is likely the protein which recognises the site will be also. Most proteins which recognise symetrical sites act as dimers and the dimer interaction leads to the symetry of the dimer. Each subunit of the dimer makes approximately 12-13 non-specific (ionic) interactions with the backbone of the DNA. The specific interactions are made by ARG 180, 185, (magenta) and GLU 181 (cyan) hydrogen bonding to the bases in the TGTGA sequence. So the protein is probably guided into place by non-specific interactions, and then "locked" in by specific ones. The cAMP (orange) bound to the CAP monomers changes the conformation of the dimer so that the distance between the recognition helices matches the spacing required for the binding. When cAMP is not bound the distance between the helices changes and the spacing is no longer optimal. The DNA is bent when the CAP protein binds, the overall bending being greater than 90o. This appears to be essential for the correct fit of the protein. It is also important in activating transcription; although it is not known whether the bending helps unwind the DNA, or position the RNA Polymerase correctly. One might speculate that the kinking of the DNA helps release the CAP protein from the DNA when the [cAMP} falls. Once the cAMP leaves the CAP the energy involved in the kinking may force the unliganded (no cAMP) protein off the DNA binding sequence. (A little bit of speculation by JWT) There are other motifs involved in DNA binding proteins, and these are dealt with in the last two tutorials on this issue. Press NEXT to view them now, or go HOME if you wish to finish for the day.
This protein has two domains: the N-terminal domain (red) which binds cAMP, and the C-terminal domain (yellow) which binds DNA. It is the latter domain which contains the helix-turn-helix motif. One helix is the recognition helix (red). It is stabilised by hydrophobic interactions with the other helix (yellow), which is almost perpendicular. The protein binds as a dimer, and dimerisation results in two recognition helices being 3.4nm apart (one turn of DNA). This is another common feature seen in many DNA binding proteins. Recognition by CAP The two-fold symmetry of the protein (after dimerisation ) once again matches the 2-fold symmetry of the target sequence. It must be noted that not ALL target sequences are symmetrical, but if they are, it is likely the protein which recognises the site will be also. Most proteins which recognise symetrical sites act as dimers and the dimer interaction leads to the symetry of the dimer. Each subunit of the dimer makes approximately 12-13 non-specific (ionic) interactions with the backbone of the DNA. The specific interactions are made by ARG 180, 185, (magenta) and GLU 181 (cyan) hydrogen bonding to the bases in the TGTGA sequence. So the protein is probably guided into place by non-specific interactions, and then "locked" in by specific ones. The cAMP (orange) bound to the CAP monomers changes the conformation of the dimer so that the distance between the recognition helices matches the spacing required for the binding. When cAMP is not bound the distance between the helices changes and the spacing is no longer optimal. The DNA is bent when the CAP protein binds, the overall bending being greater than 90o. This appears to be essential for the correct fit of the protein. It is also important in activating transcription; although it is not known whether the bending helps unwind the DNA, or position the RNA Polymerase correctly. One might speculate that the kinking of the DNA helps release the CAP protein from the DNA when the [cAMP} falls. Once the cAMP leaves the CAP the energy involved in the kinking may force the unliganded (no cAMP) protein off the DNA binding sequence. (A little bit of speculation by JWT) There are other motifs involved in DNA binding proteins, and these are dealt with in the last two tutorials on this issue. Press NEXT to view them now, or go HOME if you wish to finish for the day.
One helix is the recognition helix (red). It is stabilised by hydrophobic interactions with the other helix (yellow), which is almost perpendicular.
The protein binds as a dimer, and dimerisation results in two recognition helices being 3.4nm apart (one turn of DNA). This is another common feature seen in many DNA binding proteins.
Recognition by CAP The two-fold symmetry of the protein (after dimerisation ) once again matches the 2-fold symmetry of the target sequence. It must be noted that not ALL target sequences are symmetrical, but if they are, it is likely the protein which recognises the site will be also. Most proteins which recognise symetrical sites act as dimers and the dimer interaction leads to the symetry of the dimer. Each subunit of the dimer makes approximately 12-13 non-specific (ionic) interactions with the backbone of the DNA. The specific interactions are made by ARG 180, 185, (magenta) and GLU 181 (cyan) hydrogen bonding to the bases in the TGTGA sequence. So the protein is probably guided into place by non-specific interactions, and then "locked" in by specific ones. The cAMP (orange) bound to the CAP monomers changes the conformation of the dimer so that the distance between the recognition helices matches the spacing required for the binding. When cAMP is not bound the distance between the helices changes and the spacing is no longer optimal. The DNA is bent when the CAP protein binds, the overall bending being greater than 90o. This appears to be essential for the correct fit of the protein. It is also important in activating transcription; although it is not known whether the bending helps unwind the DNA, or position the RNA Polymerase correctly. One might speculate that the kinking of the DNA helps release the CAP protein from the DNA when the [cAMP} falls. Once the cAMP leaves the CAP the energy involved in the kinking may force the unliganded (no cAMP) protein off the DNA binding sequence. (A little bit of speculation by JWT) There are other motifs involved in DNA binding proteins, and these are dealt with in the last two tutorials on this issue. Press NEXT to view them now, or go HOME if you wish to finish for the day.
The two-fold symmetry of the protein (after dimerisation ) once again matches the 2-fold symmetry of the target sequence. It must be noted that not ALL target sequences are symmetrical, but if they are, it is likely the protein which recognises the site will be also. Most proteins which recognise symetrical sites act as dimers and the dimer interaction leads to the symetry of the dimer.
Each subunit of the dimer makes approximately 12-13 non-specific (ionic) interactions with the backbone of the DNA. The specific interactions are made by ARG 180, 185, (magenta) and GLU 181 (cyan) hydrogen bonding to the bases in the TGTGA sequence. So the protein is probably guided into place by non-specific interactions, and then "locked" in by specific ones. The cAMP (orange) bound to the CAP monomers changes the conformation of the dimer so that the distance between the recognition helices matches the spacing required for the binding. When cAMP is not bound the distance between the helices changes and the spacing is no longer optimal. The DNA is bent when the CAP protein binds, the overall bending being greater than 90o. This appears to be essential for the correct fit of the protein. It is also important in activating transcription; although it is not known whether the bending helps unwind the DNA, or position the RNA Polymerase correctly. One might speculate that the kinking of the DNA helps release the CAP protein from the DNA when the [cAMP} falls. Once the cAMP leaves the CAP the energy involved in the kinking may force the unliganded (no cAMP) protein off the DNA binding sequence. (A little bit of speculation by JWT) There are other motifs involved in DNA binding proteins, and these are dealt with in the last two tutorials on this issue. Press NEXT to view them now, or go HOME if you wish to finish for the day.
The specific interactions are made by ARG 180, 185, (magenta) and GLU 181 (cyan) hydrogen bonding to the bases in the TGTGA sequence. So the protein is probably guided into place by non-specific interactions, and then "locked" in by specific ones. The cAMP (orange) bound to the CAP monomers changes the conformation of the dimer so that the distance between the recognition helices matches the spacing required for the binding. When cAMP is not bound the distance between the helices changes and the spacing is no longer optimal.
The DNA is bent when the CAP protein binds, the overall bending being greater than 90o. This appears to be essential for the correct fit of the protein.
One might speculate that the kinking of the DNA helps release the CAP protein from the DNA when the [cAMP} falls. Once the cAMP leaves the CAP the energy involved in the kinking may force the unliganded (no cAMP) protein off the DNA binding sequence. (A little bit of speculation by JWT)
There are other motifs involved in DNA binding proteins, and these are dealt with in the last two tutorials on this issue. Press NEXT to view them now, or go HOME if you wish to finish for the day.
