Mapping Simocyclinone D8 Interaction with DNA Gyrase: Evidence for a New Binding Site on GyrB
Mapping Simocyclinone D8 Interaction with DNA Gyrase: Evidence for a New Binding Site on GyrB
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DOI：
10.1128/aac.00972-09
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发表时间：
2010-01-01
影响因子：
4.9
通讯作者：
Palumbo, M.
中科院分区：
文献类型：
--
作者：
Sissi, C.;Vazquez, E.;Palumbo, M.
Simocyclinone D8 是一种从抗生素链霉菌 Tu 6040 中分离出来的香豆素衍生物，是一种有趣的新型抗增殖剂。最初认为该药物可识别 GyrA 亚基并通过阻止其与 DNA 结合来干扰旋转酶催化循环。为了进一步表征这种抗生素的作用模式，我们通过进行蛋白质熔解和蛋白水解消化研究以及抑制测定，研究了其与重组 DNA 旋转酶 (A(2)B(2)) 及其 GyrA 和 GyrB 亚基以及这些蛋白质的各个结构域的结合。鉴定出两个结合位点，一个（预期）位于 GyrA (GyrA59) 的 N 端结构域，另一个（意外）位于 GyrB (GyrB47) 的 C 端结构域。 A 亚基的稳定似乎比 B 亚基的稳定更有效。我们的数据表明这两个不同的位点可以在重组酶中合作。
Simocyclinone D8, a coumarin derivative isolated from Streptomyces antibioticus Tu 6040, represents an interesting new antiproliferative agent. It was originally suggested that this drug recognizes the GyrA subunit and interferes with the gyrase catalytic cycle by preventing its binding to DNA. To further characterize the mode of action of this antibiotic, we investigated its binding to the reconstituted DNA gyrase (A(2)B(2)) as well as to its GyrA and GyrB subunits and the individual domains of these proteins, by performing protein melting and proteolytic digestion studies as well as inhibition assays. Two binding sites were identified, one (anticipated) in the N-terminal domain of GyrA (GyrA59) and the other (unexpected) at the C-terminal domain of GyrB (GyrB47). Stabilization of the A subunit appears to be considerably more effective than stabilization of the B subunit. Our data suggest that these two distinct sites could cooperate in the reconstituted enzyme.