Department of Biochemistry and Molecular Biophysics, Washington University School of Medicine, St Louis, MO 63110, USA.
EMBO J. 2010 Nov 17;29(22):3826-39. doi: 10.1038/emboj.2010.242. Epub 2010 Sep 28.
Escherichia coli UvrD is a 3'-5' superfamily 1A helicase/translocase involved in a variety of DNA metabolic processes. UvrD can function either as a helicase or only as an single-stranded DNA (ssDNA) translocase. The switch between these activities is controlled in vitro by the UvrD oligomeric state; a monomer has ssDNA translocase activity, whereas at least a dimer is needed for helicase activity. Although a 3'-ssDNA partial duplex provides a high-affinity site for a UvrD monomer, here we show that a monomer also binds with specificity to DNA junctions possessing a 5'-ssDNA flanking region and can initiate translocation from this site. Thus, a 5'-ss-duplex DNA junction can serve as a high-affinity loading site for the monomeric UvrD translocase, whereas a 3'-ss-duplex DNA junction inhibits both translocase and helicase activity of the UvrD monomer. Furthermore, the 2B subdomain of UvrD is important for this junction specificity. This highlights a separation of helicase and translocase function for UvrD and suggests that a monomeric UvrD translocase can be loaded at a 5'-ssDNA junction when translocation activity alone is needed.
大肠杆菌 UvrD 是一种 3'-5' 超家族 1A 解旋酶/移位酶，参与多种 DNA 代谢过程。UvrD 可以作为解旋酶或仅作为单链 DNA（ssDNA）移位酶发挥作用。这些活性之间的转换在体外受 UvrD 寡聚状态控制；单体具有 ssDNA 移位酶活性，而至少二聚体是解旋酶活性所必需的。尽管 3'-ssDNA 部分双链体为 UvrD 单体提供了高亲和力结合位点，但在这里我们表明单体也特异性地结合具有 5'-ssDNA 侧翼区的 DNA 连接点，并可以从该位点起始移位。因此，5'-ss- 双链 DNA 连接点可以作为单体 UvrD 移位酶的高亲和力加载位点，而 3'-ss- 双链 DNA 连接点抑制 UvrD 单体的移位酶和解旋酶活性。此外，UvrD 的 2B 亚结构域对于这种连接点特异性很重要。这突出了 UvrD 的解旋酶和移位酶功能的分离，并表明当仅需要移位酶活性时，单体 UvrD 移位酶可以在 5'-ssDNA 连接点加载。
