Bishop P, Jones C, Ghosh I, Chmielewski J
Department of Chemistry, Purdue University, West Lafayette, Indiana, USA.
Int J Pept Protein Res. 1995 Aug;46(2):149-54. doi: 10.1111/j.1399-3011.1995.tb01330.x.
The basic-helix-loop-helix (bHLH) region of the immunoglobulin enhancer binding protein E47 (IEB E47) was prepared in high yield by a solid-phase peptide synthesis methodology. Size-exclusion chromatography, sedimentation equilibrium and cross-linking data showed that the synthetic bHLH protein, 1, was dimeric, and higher-order aggregates of trimer and tetramer were also observed. The circular dichroism spectrum of 1 showed a high helical content, which increased upon addition of DNA containing the kappa E2 sequence. Gel mobility shift experiments showed that protein 1 bound sequence specifically to the kappa E2 sequence with a binding constant of 10(-10) M2, and had an affinity for other E box sequences as well. Comparisons between the co-crystal structure of IEB E47 with DNA and structural studies in solution showed lower helical contents in solution as would have been predicted from the crystal structure.
通过固相肽合成方法高产率地制备了免疫球蛋白增强子结合蛋白E47（IEB E47）的基本螺旋-环-螺旋（bHLH）区域。尺寸排阻色谱、沉降平衡和交联数据表明，合成的bHLH蛋白1是二聚体，还观察到三聚体和四聚体的高阶聚集体。1的圆二色光谱显示出高螺旋含量，在加入含κE2序列的DNA后增加。凝胶迁移率变动实验表明，蛋白1以10^(-10) M²的结合常数与κE2序列特异性结合，并且对其他E盒序列也有亲和力。IEB E47与DNA的共晶体结构与溶液中的结构研究之间的比较表明，溶液中的螺旋含量低于晶体结构所预测的含量。
