Cyclic diguanylate is a ubiquitous signaling molecule in bacteria: Insights into biochemistry of the GGDEF protein domain
Cyclic diguanylate is a ubiquitous signaling molecule in bacteria: Insights into biochemistry of the GGDEF protein domain
复制标题
DOI：
10.1128/jb.187.5.1792-1798.2005
复制
发表时间：
2005-03-01
影响因子：
3.2
通讯作者：
Gomelsky, M
中科院分区：
文献类型：
--
作者：
Ryjenkov, DA;Tarutina, M;Gomelsky, M
含有GGDEF结构域的蛋白质在大多数测序的细菌基因组中编码。在一些物种中，这些蛋白质与胞外多糖的生物合成、生物膜的形成、固着生活方式的建立、表面运动和基因表达的调节有关。然而，只有少数GGDEF结构域蛋白的生物化学活性已被测试。这些蛋白质被证明参与环状双（3 '-> 5'）二聚GMP（c-di-GMP）的合成或水解或环AMP的水解。为了研究细菌中GGDEF结构域的特异性，从细菌系统发育树的不同分支的随机选择的代表中，即，Thermotoga，Deinococcus-Thermus，蓝细菌，螺旋体，以及变形菌的α和γ分裂，被克隆和过表达。所有的重组蛋白进行纯化，并发现具有二鸟苷酸环化酶（DGC）的活性参与c-di-GMP的合成。来自两种蛋白质的单个GGDEF结构域被过表达、纯化，并且显示具有低水平的DGC活性。全长蛋白和单个GGDEF结构域的寡聚状态相似。这表明GGDEF结构域足以编码DGC活性;然而，酶活性受到相邻感觉蛋白结构域的高度调节。结果表明，来自伯氏疏螺旋体的GGDEF结构域蛋白Rrp 1的DGC活性严格依赖于其输入接收器结构域的磷酸化状态。本研究确定大多数GGDEF结构域蛋白是c-di-GMP特异性的，c-di-GMP合成是细菌中广泛存在的现象，并且它受到高度调节。
Proteins containing GGDEF domains are encoded in the majority of sequenced bacterial genomes. In several species, these proteins have been implicated in biosynthesis of exopolysaccharides, formation of biofilms, establishment of a sessile lifestyle, surface motility, and regulation of gene expression. However, biochemical activities of only a few GGDEF domain proteins have been tested. These proteins were shown to be involved in either synthesis or hydrolysis of cyclic-bis(3'-->5') dimeric GMP (c-di-GMP) or in hydrolysis of cyclic AMP. To investigate specificity of the GGDEF domains in Bacteria, six GGDEF domain-encoding genes from randomly chosen representatives of diverse branches of the bacterial phylogenetic tree, i.e., Thermotoga, Deinococcus-Thermus, Cyanobacteria, spirochetes, and alpha and gamma divisions of the Proteobacteria, were cloned and overexpressed. All recombinant proteins were purified and found to possess diguanylate cyclase (DGC) activity involved in c-di-GMP synthesis. The individual GGDEF domains from two proteins were overexpressed, purified, and shown to possess a low level of DGC activity. The oligomeric states of full-length proteins and individual GGDEF domains were similar. This suggests that GGDEF domains are sufficient to encode DGC activity; however, enzymatic activity is highly regulated by the adjacent sensory protein domains. It is shown that DGC activity of the GGDEF domain protein Rrp1 from Borrelia burgdorferi is strictly dependent on phosphorylation status of its input receiver domain. This study establishes that majority of GGDEF domain proteins are c-di-GMP specific, that c-di-GMP synthesis is a wide-spread phenomenon in Bacteria, and that it is highly regulated.