DNA促旋酶,一种普遍存在的细菌酶,是通过2个GyrA亚基和2个GyrB亚基的异源四聚形成的IIA型拓扑异构酶,形成活性复合体。DNA促旋酶可以在GyrA的C末端结构域(CTD)周围形成DNA环,并使一个DNA双链体通过在另一个双链体中建立的瞬时双链断裂(DSB)。这导致从正(1)到负(-1)的超线圈的转换,从而通过步骤2将负超螺旋引入细菌基因组,这是DNA复制和转录所必需的活性。GyrA二聚体中的强蛋白质界面必须被破坏以允许运输的DNA片段通过,并且通常认为该界面通常是稳定的,并且仅在运输DNA时才打开。防止在基因组中引入有害的DSB。在本文中,我们表明DNA促旋酶可以在两个活性异四聚体之间交换其DNA切割界面。这种所谓的接口\'交换\'(IS)可以在解决方案中的几分钟内发生。我们还表明,旋转酶对DNA的弯曲对于切割至关重要,但对于DNA结合本身并不重要,并且有利于IS。DNA包裹和过量的GyrB也有利于界面交换。我们建议接近,通过GyrB寡聚化以及沿着DNA长度的结合和包裹促进,在两个异四聚体之间有利于快速界面交换。这种交换不需要ATP,发生在氟喹诺酮类药物的存在下,并提高了仅通过促旋酶活性进行非同源重组的可能性。促旋酶进行界面交换的能力解释了促旋酶异二聚体,含有单个活性位点酪氨酸,可以进行双链通道反应,因此对最近提出的DNA旋转酶的“旋转”机制提出了另一种解释(Gubaev等人。,2016)。
DNA gyrase, a ubiquitous bacterial enzyme, is a type IIA topoisomerase formed by heterotetramerisation of 2 GyrA subunits and 2 GyrB subunits, to form the active complex. DNA gyrase can loop DNA around the C-terminal domains (CTDs) of GyrA and pass one DNA duplex through a transient double-strand break (DSB) established in another duplex. This results in the conversion from a positive (+1) to a negative (-1) supercoil, thereby introducing negative supercoiling into the bacterial genome by steps of 2, an activity essential for DNA replication and transcription. The strong protein interface in the GyrA dimer must be broken to allow passage of the transported DNA segment and it is generally assumed that the interface is usually stable and only opens when DNA is transported, to prevent the introduction of deleterious DSBs in the genome. In this paper, we show that DNA gyrase can exchange its DNA-cleaving interfaces between two active heterotetramers. This so-called interface \'swapping\' (IS) can occur within a few minutes in solution. We also show that bending of DNA by gyrase is essential for cleavage but not for DNA binding per se and favors IS. Interface swapping is also favored by DNA wrapping and an excess of GyrB. We suggest that proximity, promoted by GyrB oligomerization and binding and wrapping along a length of DNA, between two heterotetramers favors rapid interface swapping. This swapping does not require ATP, occurs in the presence of fluoroquinolones, and raises the possibility of non-homologous recombination solely through gyrase activity. The ability of gyrase to undergo interface swapping explains how gyrase heterodimers, containing a single active-site tyrosine, can carry out double-strand passage reactions and therefore suggests an alternative explanation to the recently proposed \'swivelling\' mechanism for DNA gyrase (Gubaev et al., 2016).