Initiation of lambda DNA replication. The Escherichia coli small heat shock proteins, DnaJ and GrpE, increase DnaK's affinity for the lambda P protein.
Initiation of lambda DNA replication. The Escherichia coli small heat shock proteins, DnaJ and GrpE, increase DnaK's affinity for the lambda P protein.
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DOI：
10.1016/s0021-9258(18)53470-0
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发表时间：
1993-03
期刊：
影响因子：
--
通讯作者：
J. Osipiuk;C. Georgopoulos;M. Żylicz
中科院分区：
文献类型：
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作者：
J. Osipiuk;C. Georgopoulos;M. Żylicz
众所周知，噬菌体lambda复制的起始需要在ori lambda DNA位点有序组装lambda O.lambda P.DnaB解旋酶蛋白前染色体复合体。DnaK、DnaJ和GrpE热休克蛋白共同作用，破坏lambda P.DnaB复合物的稳定性，从而释放DnaB，并允许它在ori lambda位点附近解开lambda DNA。分解反应的第一步是DnaK与P蛋白的结合。在本报告中，我们研究了DnaJ和GrpE蛋白对lambda P.DnaK复合物稳定性的影响。我们提出了以下蛋白质-蛋白质复合物存在的证据：lambda P.DnaK, lambda P.DnaJ, DnaJ。DnaK DnaK。P.DnaK.GrpE。我们的研究结果表明，GrpE单独存在会破坏lambda P.DnaK复合物的稳定性，而DnaJ单独存在则会稳定lambda P.DnaK复合物。通过免疫沉淀，我们发现在GrpE存在的情况下，DnaK对lambda P.DnaJ复合物的亲和力比单独存在时更高。通过与戊二醛交联，我们发现DnaK的低聚形式比单体DnaK对λ P具有更高的亲和力。然而，在GrpE存在下，单体DnaK可以有效地结合lambda P蛋白。这些发现有助于解释我们之前的结果，即在依赖grpe的lambda DNA复制系统中，DnaK蛋白的需求可以减少10倍。
It is known that the initiation of bacteriophage lambda replication requires the orderly assembly of the lambda O.lambda P.DnaB helicase protein preprimosomal complex at the ori lambda DNA site. The DnaK, DnaJ, and GrpE heat shock proteins act together to destabilize the lambda P.DnaB complex, thus freeing DnaB and allowing it to unwind lambda DNA near the ori lambda site. The first step of this disassembly reaction is the binding of DnaK to the lambda P protein. In this report, we examined the influence of the DnaJ and GrpE proteins on the stability of the lambda P.DnaK complex. We present evidence for the existence of the following protein-protein complexes: lambda P.DnaK, lambda P.DnaJ, DnaJ.DnaK, DnaK.GrpE, and lambda P.DnaK.GrpE. Our results suggest that the presence of GrpE alone destabilizes the lambda P.DnaK complex, whereas the presence of DnaJ alone stabilizes the lambda P.DnaK complex. Using immunoprecipitation, we show that in the presence of GrpE, DnaK exhibits a higher affinity for the lambda P.DnaJ complex than it does alone. Using cross-linking with glutaraldehyde, we show that oligomeric forms of DnaK exhibit a higher affinity for lambda P than monomeric DnaK. However, in the presence of GrpE, monomeric DnaK can efficiently bind lambda P protein. These findings help explain our previous results, namely that in the GrpE-dependent lambda DNA replication system, the DnaK protein requirement can be reduced up to 10-fold.