May Felicity E B, Church Stephen T, Major Sarah, Westley Bruce R
Department of Pathology, School of Clinical and Laboratory Sciences, University of Newcastle upon Tyne, Royal Victoria Infirmary, Newcastle upon Tyne NE1 4LP, UK.
Biochemistry. 2003 Jul 15;42(27):8250-9. doi: 10.1021/bi030025l.
The human trefoil proteins TFF1 and TFF3 are expressed predominantly in the gastrointestinal tract. They are also expressed and regulated by estrogens in malignant breast epithelial cells. TFF1 and TFF3 are small cysteine-rich acidic secreted proteins of 60 and 59 amino acids with similar isoelectric points of 4.75 and 3.94, respectively. Each contains one trefoil domain that is characterized by several conserved features including six cysteine residues with conserved spacing. TFF1 and TFF3 form intermolecular disulfide bonds via an extra-trefoil domain cysteine residue and are present in vivo as monomers and homodimers and as complexes with other proteins. The TFF1 dimer is more active than the TFF1 monomer. In the present study the hydrodynamic and charge properties of TFF1 and TFF3 monomers and homodimers have been compared and shown to differ markedly. Notably, TFF1 is significantly more asymmetric than TFF3 (frictional coefficients 1.25 and 1.12, respectively, p < 0.001), and homodimerization of TFF1 results in a greater increase in asymmetry than for TFF3. The overall charges of TFF1 and TFF3 are very different at neutral pH. Titration curves predicted significant differences in charge across a wide pH range that agreed well with experimental data. The locations of charged amino acids in the primary sequences and in the tertiary structures of TFF1 and TFF3 were examined. This revealed interesting divergence in both the distribution and local topology of charged amino acid side chains. The significant differences between the shape, size, and surface charge of these two closely related molecules may account for their divergent biological activities.
人三叶因子蛋白TFF1和TFF3主要在胃肠道表达。它们也在恶性乳腺上皮细胞中表达并受雌激素调节。TFF1和TFF3是富含半胱氨酸的小酸性分泌蛋白，分别由60和59个氨基酸组成，等电点分别为4.75和3.94。每个蛋白都含有一个三叶结构域，其特征是具有几个保守特征，包括六个间距保守的半胱氨酸残基。TFF1和TFF3通过三叶结构域以外的半胱氨酸残基形成分子间二硫键，在体内以单体、同二聚体以及与其他蛋白质的复合物形式存在。TFF1二聚体比TFF1单体更具活性。在本研究中，对TFF1和TFF3单体及同二聚体的流体动力学和电荷特性进行了比较，结果显示它们有显著差异。值得注意的是，TFF1的不对称性明显高于TFF3（摩擦系数分别为1.25和1.12，p<0.001），并且TFF1同二聚化导致的不对称性增加比TFF3更大。在中性pH条件下，TFF1和TFF3的总电荷差异很大。滴定曲线预测在很宽的pH范围内电荷有显著差异，这与实验数据非常吻合。研究了TFF1和TFF3一级序列和三级结构中带电荷氨基酸的位置。这揭示了带电荷氨基酸侧链在分布和局部拓扑结构上有趣的差异。这两个密切相关分子在形状、大小和表面电荷上的显著差异可能解释了它们不同的生物学活性。
