Bajorath J, Marken J S, Chalupny N J, Spoon T L, Siadak A W, Gordon M, Noelle R J, Hollenbaugh D, Aruffo A
Bristol-Myers Squibb Pharmaceutical Research Institute, Seattle, Washington 98121, USA.
Biochemistry. 1995 Aug 8;34(31):9884-92. doi: 10.1021/bi00031a009.
The interaction between gp39 (CD40L, TRAP, T-BAM) on activated T cells and mast cells and CD40 on antigen-presenting cells modulates immune responses. Gp39 and CD40 are homologous to tumor necrosis factor (TNF) and its receptor (TNFR), respectively. The TNF-beta/TNFR interaction has been analyzed on the basis of mutagenesis experiments and crystal structures. Using the interaction of TNF-beta/TNFR as a guide, we previously reported a site-directed mutagenesis study in which we identified residues in gp39 (K143, Y145) and CD40 (Y82, D84, N86) involved in gp39/CD40 interactions. Here we describe the use of the TNF-beta/TNFR complex crystal structure as a template to prepare molecular models of gp39, CD40, and their approximate interaction. The application of these models has allowed us to extend our mutagenesis analysis of gp39/CD40 interactions. These experiments have led to the identification of additional gp39 (Y146, R203, Q220) and CD40 (E74, E117) residues that contribute to the gp39/CD40 interaction. We also further explored the importance of gp39 residue Y145 and CD40 residue Y82 for the gp39/CD40 interaction by conservatively replacing these residues with Phe. The results of these studies have enabled us to approximately outline the binding sites in gp39 and CD40. It appears that the gp39/CD40 interaction is centered on at least two clusters of residues and involves residues of two adjacent gp39 monomers. The molecular regions involved in the gp39/CD40 interaction essentially correspond to those in the homologous TNF-beta/TNFR system.
活化T细胞上的gp39（CD40L、TRAP、T-BAM）与肥大细胞以及抗原呈递细胞上的CD40之间的相互作用可调节免疫反应。Gp39和CD40分别与肿瘤坏死因子（TNF）及其受体（TNFR）同源。基于诱变实验和晶体结构对TNF-β/TNFR的相互作用进行了分析。以TNF-β/TNFR的相互作用为指导，我们之前报道了一项定点诱变研究，其中我们确定了gp39（K143、Y145）和CD40（Y82、D84、N86）中参与gp39/CD40相互作用的残基。在此我们描述了以TNF-β/TNFR复合晶体结构为模板来制备gp39、CD40及其近似相互作用的分子模型。这些模型的应用使我们能够扩展对gp39/CD40相互作用的诱变分析。这些实验导致鉴定出了有助于gp39/CD40相互作用的其他gp39（Y146、R203、Q220）和CD40（E74、E117）残基。我们还通过用苯丙氨酸保守取代这些残基，进一步探究了gp39残基Y145和CD40残基Y82对gp39/CD40相互作用的重要性。这些研究结果使我们能够大致勾勒出gp39和CD40中的结合位点。看来gp39/CD40相互作用至少集中在两个残基簇上，并且涉及两个相邻gp39单体的残基。gp39/CD40相互作用中涉及的分子区域基本上与同源TNF-β/TNFR系统中的区域相对应。
