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Thermodynamics of DNA target site recognition by homing endonucleases.

Eastberg JH, McConnell Smith A, Zhao L, Ashworth J, Shen BW, Stoddard BL - Nucleic Acids Res. (2007)

Bottom Line: While the balance of DeltaH and TDeltaS are not strongly correlated with the overall extent of DNA bending, unfavorable DeltaH(binding) is associated with unstacking of individual base steps in the target site.The effects of deleterious basepair substitutions in the optimal target sites of two LAGLIDADG homing endonucleases, and the subsequent effect of redesigning one of those endonucleases to accommodate that DNA sequence change, were also measured.The substitution of base-specific hydrogen bonds in a wild-type endonuclease/DNA complex with hydrophobic van der Waals contacts in a redesigned complex reduced the ability to discriminate between sites, due to nonspecific DeltaS(binding).

View Article: PubMed Central - PubMed

Affiliation: Division of Basic Sciences, Fred Hutchinson Cancer Research Center, 1100 Fairview Avenue North, A3-025 Seattle, WA 98109, USA.

ABSTRACT
The thermodynamic profiles of target site recognition have been surveyed for homing endonucleases from various structural families. Similar to DNA-binding proteins that recognize shorter target sites, homing endonucleases display a narrow range of binding free energies and affinities, mediated by structural interactions that balance the magnitude of enthalpic and entropic forces. While the balance of DeltaH and TDeltaS are not strongly correlated with the overall extent of DNA bending, unfavorable DeltaH(binding) is associated with unstacking of individual base steps in the target site. The effects of deleterious basepair substitutions in the optimal target sites of two LAGLIDADG homing endonucleases, and the subsequent effect of redesigning one of those endonucleases to accommodate that DNA sequence change, were also measured. The substitution of base-specific hydrogen bonds in a wild-type endonuclease/DNA complex with hydrophobic van der Waals contacts in a redesigned complex reduced the ability to discriminate between sites, due to nonspecific DeltaS(binding).

Show MeSH
Isothermal enthalpy–entropy compensation by homing endonucleases and other DNA-binding proteins. The enthalpic (ΔH) and entropic (−TΔS) contributions to site-specific DNA recognition of different protein–DNA complexes, including several representative wild-type homing endonucleases (highlighted in red) are shown. The thermodynamic values for previously studied DNA-binding proteins are shown in blue, and are taken from previous analyses by Jen-Jacobson et al. (1) and references therein.
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Figure 3: Isothermal enthalpy–entropy compensation by homing endonucleases and other DNA-binding proteins. The enthalpic (ΔH) and entropic (−TΔS) contributions to site-specific DNA recognition of different protein–DNA complexes, including several representative wild-type homing endonucleases (highlighted in red) are shown. The thermodynamic values for previously studied DNA-binding proteins are shown in blue, and are taken from previous analyses by Jen-Jacobson et al. (1) and references therein.

Mentions: The binding of 11 separate combinations of homing endonucleases and various DNA target sites, representing five major structural classes of these proteins, was analyzed by ITC. The DNA constructs used for these studies are shown in Table 1; the structure of each wild-type protein/DNA complex is shown in Figure 1. The affinities and thermodynamic signatures of these interactions are summarized in Table 2 and Figures 2 and 3.Figure 2.


Thermodynamics of DNA target site recognition by homing endonucleases.

Eastberg JH, McConnell Smith A, Zhao L, Ashworth J, Shen BW, Stoddard BL - Nucleic Acids Res. (2007)

Isothermal enthalpy–entropy compensation by homing endonucleases and other DNA-binding proteins. The enthalpic (ΔH) and entropic (−TΔS) contributions to site-specific DNA recognition of different protein–DNA complexes, including several representative wild-type homing endonucleases (highlighted in red) are shown. The thermodynamic values for previously studied DNA-binding proteins are shown in blue, and are taken from previous analyses by Jen-Jacobson et al. (1) and references therein.
© Copyright Policy - creative-commons
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2175346&req=5

Figure 3: Isothermal enthalpy–entropy compensation by homing endonucleases and other DNA-binding proteins. The enthalpic (ΔH) and entropic (−TΔS) contributions to site-specific DNA recognition of different protein–DNA complexes, including several representative wild-type homing endonucleases (highlighted in red) are shown. The thermodynamic values for previously studied DNA-binding proteins are shown in blue, and are taken from previous analyses by Jen-Jacobson et al. (1) and references therein.
Mentions: The binding of 11 separate combinations of homing endonucleases and various DNA target sites, representing five major structural classes of these proteins, was analyzed by ITC. The DNA constructs used for these studies are shown in Table 1; the structure of each wild-type protein/DNA complex is shown in Figure 1. The affinities and thermodynamic signatures of these interactions are summarized in Table 2 and Figures 2 and 3.Figure 2.

Bottom Line: While the balance of DeltaH and TDeltaS are not strongly correlated with the overall extent of DNA bending, unfavorable DeltaH(binding) is associated with unstacking of individual base steps in the target site.The effects of deleterious basepair substitutions in the optimal target sites of two LAGLIDADG homing endonucleases, and the subsequent effect of redesigning one of those endonucleases to accommodate that DNA sequence change, were also measured.The substitution of base-specific hydrogen bonds in a wild-type endonuclease/DNA complex with hydrophobic van der Waals contacts in a redesigned complex reduced the ability to discriminate between sites, due to nonspecific DeltaS(binding).

View Article: PubMed Central - PubMed

Affiliation: Division of Basic Sciences, Fred Hutchinson Cancer Research Center, 1100 Fairview Avenue North, A3-025 Seattle, WA 98109, USA.

ABSTRACT
The thermodynamic profiles of target site recognition have been surveyed for homing endonucleases from various structural families. Similar to DNA-binding proteins that recognize shorter target sites, homing endonucleases display a narrow range of binding free energies and affinities, mediated by structural interactions that balance the magnitude of enthalpic and entropic forces. While the balance of DeltaH and TDeltaS are not strongly correlated with the overall extent of DNA bending, unfavorable DeltaH(binding) is associated with unstacking of individual base steps in the target site. The effects of deleterious basepair substitutions in the optimal target sites of two LAGLIDADG homing endonucleases, and the subsequent effect of redesigning one of those endonucleases to accommodate that DNA sequence change, were also measured. The substitution of base-specific hydrogen bonds in a wild-type endonuclease/DNA complex with hydrophobic van der Waals contacts in a redesigned complex reduced the ability to discriminate between sites, due to nonspecific DeltaS(binding).

Show MeSH