Stokes H W, O'Gorman D B, Recchia G D, Parsekhian M, Hall R M
School of Biological Sciences, Macquarie University Sydney, NSW, Australia.
Mol Microbiol. 1997 Nov;26(4):731-45. doi: 10.1046/j.1365-2958.1997.6091980.x.
The integration of gene cassettes into integrons is effected by site-specific recombination catalysed by an integrase, IntI, encoded by the integron. The cassette-associated recombination sites, 59-base elements, are not highly conserved and vary in length from 57 to 141 bp. They can be identified by their location and the relationship of over 20 bp at their outer ends to consensus sequences that are imperfect inverted repeats of one another. The recombination cross-over occurs close to one end of the 59-base element, within a conserved core site with the consensus sequence GTTAGGC or GTTRRRY. By introducing single-base changes at each of these positions in the aadB 59-base element, bases that are critical for site activity were identified. The recombination cross-over was also localized to a unique position between the adjacent G and T residues. Changes introduced in the conserved AAC of the inverse core site (GCCTAAC or RYYYAAC) located at the opposite end of the 59-base element also reduced site activity but to a lesser extent. Sequences of rare recombinants revealed an alternative position for strand exchange and led to the conclusion that 59-base elements comprise two simple sites, analogous to those recognized by other integrases, with each simple site made up of a pair of inversely oriented IntI binding domains separated by a spacer of 7 or 8 bp. Re-examination of the sequences of all known 59-base elements revealed that this simple site configuration was present at both the left and right ends in all 59-base elements. The identity of bases in the spacer is not required for efficient recombination and the cross-over is located at one end of the spacer, suggesting that during IntI1-mediated recombination only one strand exchange occurs.
基因盒整合到整合子中是由整合子编码的整合酶IntI催化的位点特异性重组实现的。与基因盒相关的重组位点，即59碱基元件，并非高度保守，长度在57至141 bp之间变化。它们可以通过其位置以及其外端超过20 bp与彼此为不完全反向重复的共有序列的关系来识别。重组交叉发生在59碱基元件一端附近，在一个具有共有序列GTTAGGC或GTTRRRY的保守核心位点内。通过在aadB 59碱基元件的每个这些位置引入单碱基变化，确定了对位点活性至关重要的碱基。重组交叉也定位在相邻G和T残基之间的一个独特位置。在位于59碱基元件另一端的反向核心位点（GCCTAAC或RYYYAAC）的保守AAC中引入的变化也降低了位点活性，但程度较小。罕见重组体的序列揭示了链交换的另一个位置，并得出结论，59碱基元件由两个简单位点组成，类似于其他整合酶识别的位点，每个简单位点由一对反向排列的IntI结合结构域组成，中间由7或8 bp的间隔区隔开。对所有已知59碱基元件序列的重新检查表明，这种简单位点结构在所有59碱基元件的左端和右端都存在。间隔区中碱基的一致性对于有效重组不是必需的，并且交叉位于间隔区的一端，这表明在IntI1介导的重组过程中只发生一次链交换。
