Electrostatic interactions during acidic phospholipid reactivation of DnaA protein, the Escherichia coli initiator of chromosomal replication.
Electrostatic interactions during acidic phospholipid reactivation of DnaA protein, the Escherichia coli initiator of chromosomal replication.
复制标题
DnaA 蛋白（大肠杆菌染色体复制的启动子）的酸性磷脂重新激活过程中的静电相互作用。
DOI：
10.1021/bi982733q
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发表时间：
1999
期刊：
影响因子：
--
通讯作者：
Crooke,E
中科院分区：
文献类型：
--
作者：
Kitchen,JL;Li,Z;Crooke,E
DnaA蛋白启动大肠杆菌染色体复制受ATP和ADP紧密结合的强烈影响。流体双层中的阴离子磷脂在体外促进无活性的ADP-DnaA蛋白转化为具有复制活性的ATP-DnaA蛋白，因此可能在调节DnaA活性中起关键作用。以前的研究表明，在这种再活化过程中，DnaA蛋白的特定区域以酸性磷脂依赖性方式插入脂质双层的疏水部分。为了阐明在再活化过程中对酸性磷脂的要求，研究了DnaA和脂质相互作用中静电力的贡献。DnaA−脂质结合需要阴离子磷脂，而DnaA−脂质结合以及脂质介导的DnaA结合核苷酸的释放都受到离子强度增加的抑制，这表明静电相互作用参与了这些过程。随着酸性磷脂的囊泡含量增加，核苷酸释放和DnaA−脂质结合以线性平行方式增加。考虑到DnaA−膜结合、DnaA插入膜以及随后的核苷酸释放都需要阴离子磷脂，酸性头基可能是将DnaA蛋白募集到膜上以插入并随后重新激活复制所必需的。
The initiation ofEscherichia colichromosomal replication by DnaA protein is strongly influenced by the tight binding of the nucleotides ATP and ADP. Anionic phospholipids in a fluid bilayer promote the conversion of inactive ADP-DnaA protein to replicatively active ATP-DnaA protein in vitro, and thus likely play a key role in regulating DnaA activity. Previous studies have revealed that, during this reactivation, a specific region of DnaA protein inserts into the hydrophobic portion of the lipid bilayer in an acidic phospholipid-dependent manner. To elucidate the requirement for acidic phospholipids in the reactivation process, the contribution of electrostatic forces in the interaction of DnaA and lipid was examined. DnaA−lipid binding required anionic phospholipids, and DnaA−lipid binding as well as lipid-mediated release of DnaA-bound nucleotide were inhibited by increased ionic strength, suggesting the involvement of electrostatic interactions in these processes. As the vesicular content of acidic phospholipids was increased, both nucleotide release and DnaA−lipid binding increased in a linear, parallel manner. Given that DnaA−membrane binding, the insertion of DnaA into the membrane, and the consequent nucleotide release all require anionic phospholipids, the acidic headgroup may be necessary to recruit DnaA protein to the membrane for insertion and subsequent reactivation for replication.