Kombo D C, Némethy G, Gibson K D, Rackovsky S, Scheraga H A
Department of Biomathematical Sciences, Mount Sinai School of Medicine, NY 10029, USA.
J Mol Biol. 1996 Mar 1;256(3):517-32. doi: 10.1006/jmbi.1996.0105.
Binding of the N-terminal domain of the lambda repressor to DNA is coupled to dimerization. Hydrophobic interactions between helix-5 and helix-5' drive the packing at the dimer interface. We have carried out computations of the conformational energy of packing of the fifth helices (and of the helix-4-loop-helix-5 portions) of variants of the lambda repressor operator binding domain, using an ECEPP/3-based packing algorithm. Here, we report the results for 26 mutants chosen among those that hve been characterized experimentally. We find that the relative orientation of the fifth helices for active mutants is very similar to the wild-type. The fifth helices of the inactive mutants have a significantly different relative orientation. This result illustrates that a unique specific orientation pattern of helix-5 relative to helix-5' is required for dimerization-coupled DNA binding activity. This finding is further supported by computational studies of the whole N-terminal domain of ten variants that showed that the active mutants, including the wild-type protein, have similar values of the number of contacts between the two monomers in the dimer, involving two amino acid residues of the fifth helices (positions 84 and 87 in each monomer). A decrease in the number of such contacts abolishes DNA-binding activity. Furthermore, all active mutants have their "DNA-recognition helices", numbers 3 and 3' positioned so that they can fit in the DNA operator like those of the wild-type protein, while some inactive mutants exhibit a substantial change in the relative orientation of their recognition helices.
λ阻遏物N端结构域与DNA的结合与二聚化相关联。螺旋5和螺旋5'之间的疏水相互作用驱动二聚体界面处的堆积。我们使用基于ECEPP/3的堆积算法，对λ阻遏物操纵子结合结构域变体的第五螺旋（以及螺旋4-环-螺旋5部分）的堆积构象能量进行了计算。在此，我们报告从已进行实验表征的突变体中挑选出的26个突变体的结果。我们发现，活性突变体的第五螺旋的相对取向与野生型非常相似。非活性突变体的第五螺旋具有明显不同的相对取向。这一结果表明，二聚化偶联的DNA结合活性需要螺旋5相对于螺旋5'的独特特定取向模式。对十个变体的整个N端结构域的计算研究进一步支持了这一发现，该研究表明，包括野生型蛋白在内的活性突变体在二聚体中两个单体之间的接触数具有相似的值，涉及第五螺旋的两个氨基酸残基（每个单体中的第84和87位）。这种接触数的减少会消除DNA结合活性。此外，所有活性突变体的“DNA识别螺旋”，即螺旋3和螺旋3'的定位方式使得它们能够像野生型蛋白那样适配于DNA操纵子，而一些非活性突变体的识别螺旋的相对取向则发生了显著变化。
