Stewart J, Hingorani M M, Kelman Z, O'Donnell M
Rockefeller University and Howard Hughes Medical Institute, Laboratory of DNA Replication, New York, New York 10021, USA.
J Biol Chem. 2001 Jun 1;276(22):19182-9. doi: 10.1074/jbc.M100592200. Epub 2001 Mar 6.
The beta sliding clamp encircles the primer-template and tethers DNA polymerase III holoenzyme to DNA for processive replication of the Escherichia coli genome. The clamp is formed via hydrophobic and ionic interactions between two semicircular beta monomers. This report demonstrates that the beta dimer is a stable closed ring and is not monomerized when the gamma complex clamp loader (gamma(3)delta(1)delta(1)chi(1)psi(1)) assembles the beta ring around DNA. delta is the subunit of the gamma complex that binds beta and opens the ring; it also does not appear to monomerize beta. Point mutations were introduced at the beta dimer interface to test its structural integrity and gain insight into its interaction with delta. Mutation of two residues at the dimer interface of beta, I272A/L273A, yields a stable beta monomer. We find that delta binds the beta monomer mutant at least 50-fold tighter than the beta dimer. These findings suggest that when delta interacts with the beta clamp, it binds one beta subunit with high affinity and utilizes some of that binding energy to perform work on the dimeric clamp, probably cracking one dimer interface open.
β滑动夹环绕引物模板，并将DNA聚合酶III全酶连接到DNA上，以实现大肠杆菌基因组的持续复制。该夹子是通过两个半圆形β单体之间的疏水和离子相互作用形成的。本报告表明，β二聚体是一个稳定的闭环，当γ复合物夹子加载器（γ(3)δ(1)δ(1)χ(1)ψ(1)）围绕DNA组装β环时，它不会单体化。δ是γ复合物中与β结合并打开环的亚基；它似乎也不会使β单体化。在β二聚体界面引入点突变，以测试其结构完整性，并深入了解其与δ的相互作用。β二聚体界面处的两个残基I272A/L273A发生突变，产生了一个稳定的β单体。我们发现，δ与β单体突变体的结合比与β二聚体的结合至少紧密50倍。这些发现表明，当δ与β夹子相互作用时，它以高亲和力结合一个β亚基，并利用部分结合能对二聚体夹子做功，可能会打开一个二聚体界面。
