Crystal structure of the tandem GAF domains from a cyanobacterial adenylyl cyclase: Modes of ligand binding and dimerization
Crystal structure of the tandem GAF domains from a cyanobacterial adenylyl cyclase: Modes of ligand binding and dimerization
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DOI：
10.1073/pnas.0409913102
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发表时间：
2005-02-22
影响因子：
11.1
通讯作者：
Linder, JU
中科院分区：
文献类型：
--
作者：
Martinez, SE;Bruder, S;Linder, JU
在一些物种中，GAF 结构域是广泛表达的调节酶活性的小分子结合结构域，已知可以结合环核苷酸。然而，对于任何 GAF 结构域，环核苷酸结合影响酶活性的分子机制尚不清楚。在蓝藻鱼腥藻中，cyaB1 和 cyaB2 基因编码腺苷酸环化酶，这些酶通过 cAMP 与其 N 末端 GAF 结构域的结合而受到刺激。用哺乳动物磷酸二酯酶 2A GAF-A/B 串联结构域替换 cyaB1 中的串联 GAF-A/B 结构域，可以通过 cGMP 调节嵌合蛋白，这表明激活机制高度保守。在这里，我们描述了结合 cAMP 的 cyaB2 串联 GAF-A/B 结构域的 1.9 埃晶体结构，并将其与先前报道的 PDE2A GAF-A/B 结构进行比较。出乎意料的是，cyaB2 GAF-A/B 二聚体是反平行的，与 PDE2A 的平行二聚体不同。此外，GAF-A和-B中都存在明显的cAMP电子密度，而在PDE2A中，cGMP仅在GAF-B中发现。磷酸基和核糖基团的接触与 PDE2A 中的相似。然而，嘌呤结合袋看起来与 PDE2A GAF-B 中的非常不同。 beta2-beta3 环的差异表明，该环赋予了该区域以及许多其他 GAF 结构域的大部分配体特异性。最后，保守的天冬酰胺似乎是 NKFDE 标志性基序的新成员，并提出了该基序稳定 cNMP 结合袋的机制。
In several species, GAF domains, which are widely expressed small-molecule-binding domains that regulate enzyme activity, are known to bind cyclic nucleotides. However, the molecular mechanism by which cyclic nucleotide binding affects enzyme activity is not known for any GAF domain. In the cyanobacterium, Anabaena, the cyaB1 and cyaB2 genes encode adenylyl cyclases that are stimulated by binding of cAMP to their N-terminal GAF domains. Replacement of the tandem GAF-A/B domains in cyaB1 with the mammalian phosphodiesterase 2A GAF-A/B tandem domains allows regulation of the chimeric protein by cGMP, suggesting a highly conserved mechanism of activation. Here, we describe the 1.9-Angstrom crystal structure of the tandem GAF-A/B domains of cyaB2 with bound cAMP and compare it to the previously reported structure of the PDE2A GAF-A/B. Unexpectedly, the cyaB2 GAF-A/B dimer is antiparallel, unlike the parallel dimer of PDE2A. Moreover, there is clear electron density for cAMP in both GAF-A and -B, whereas in PDE2A, cGMP is found only in GAF-B. Phosphate and ribose group contacts are similar to those in PDE2A. However, the purine-binding pockets appear very different from that in PDE2A GAF-B. Differences in the beta2-beta3 loop suggest that this loop confers much of the ligand specificity in this and perhaps in many other GAF domains. Finally, a conserved asparagine appears to be a new addition to the signature NKFDE motif, and a mechanism for this motif to stabilize the cNMP-binding pocket is proposed.