McDonald Robert J, Kahn Jason D, Maher L James
Medical Scientist Training Program, Mayo Clinic College of Medicine, Rochester, MN 55905, USA.
Nucleic Acids Res. 2006;34(17):4846-56. doi: 10.1093/nar/gkl552. Epub 2006 Sep 14.
We wish to understand the role of electrostatics in DNA stiffness and bending. The DNA charge collapse model suggests that mutual electrostatic repulsions between neighboring phosphates significantly contribute to DNA stiffness. According to this model, placement of fixed charges near the negatively charged DNA surface should induce bending through asymmetric reduction or enhancement of these inter-phosphate repulsive forces. We have reported previously that charged variants of the elongated basic-leucine zipper (bZIP) domain of Gcn4p bend DNA in a manner consistent with this charge collapse model. To extend this result to a more globular protein, we present an investigation of the dimeric basic-helix-loop-helix (bHLH) domain of Pho4p. The 62 amino acid bHLH domain has been modified to position charged amino acid residues near one face of the DNA double helix. As observed for bZIP charge variants, DNA bending toward appended cations (away from the protein:DNA interface) is observed. However, unlike bZIP proteins, DNA is not bent away from bHLH anionic charges. This finding can be explained by the structure of the more globular bHLH domain which, in contrast to bZIP proteins, makes extensive DNA contacts along the binding face.
我们希望了解静电在DNA刚性和弯曲中的作用。DNA电荷坍塌模型表明，相邻磷酸基团之间的相互静电排斥对DNA刚性有显著贡献。根据该模型，在带负电荷的DNA表面附近放置固定电荷应通过不对称地降低或增强这些磷酸间排斥力来诱导弯曲。我们之前报道过，Gcn4p的延伸碱性亮氨酸拉链（bZIP）结构域的带电变体以与该电荷坍塌模型一致的方式使DNA弯曲。为了将这一结果扩展到更球状的蛋白质，我们对Pho4p的二聚体碱性螺旋-环-螺旋（bHLH）结构域进行了研究。62个氨基酸的bHLH结构域已被修饰，以便在DNA双螺旋的一个面附近定位带电荷的氨基酸残基。正如在bZIP电荷变体中观察到的那样，观察到DNA向附加阳离子弯曲（远离蛋白质：DNA界面）。然而，与bZIP蛋白不同的是，DNA不会从bHLH阴离子电荷处弯曲。这一发现可以用更球状的bHLH结构域的结构来解释，与bZIP蛋白相比，该结构域沿着结合面与DNA有广泛的接触。
