Senecoff J F, Cox M M
J Biol Chem. 1986 Jun 5;261(16):7380-6.
The 2 mu plasmid of the yeast Saccharomyces cerevisiae encodes a site-specific recombination system consisting of plasmid-encoded FLP protein and two recombination sites on the plasmid. The recombination site possesses a specific orientation, which is determined by an asymmetric 8-base pair spacer sequence separating two 13-base pair inverted repeats. The outcome or directionality of site-specific recombination is defined by the alignment of two sites in the same orientation during the reaction. Sites containing point mutations or 1-base pair insertions or deletions within the spacer generally undergo recombination with unaltered sites at reduced levels. In contrast, recombination between the two identical mutant sites (where homology is restored) proceeds efficiently in all cases. Sites containing spacer sequences of 10 base pairs or more are nonfunctional under all conditions. A recombination site in which 5 base pairs are changed to yield an entirely symmetrical spacer sequence again recombines efficiently, but only with an identical site. This reaction, in addition, produces a variety of new products which can only result from random alignment of the two sites undergoing recombination, i.e. the reaction no longer exhibits directionality. These and other results demonstrate that both the efficiency and directionality of site-specific recombination is dependent upon homology between spacer sequences of the two recombining sites. This further implies that critical DNA-DNA interactions between the spacer region of the two sites involved in the reaction occur at some stage during site-specific recombination in this system. The specific spacer sequence itself appears to be unimportant as long as homology is maintained; thus, these sequences are probably not involved in recognition by FLP protein.
酿酒酵母的2μm质粒编码一种位点特异性重组系统，该系统由质粒编码的FLP蛋白和质粒上的两个重组位点组成。重组位点具有特定的方向，这由一个不对称的8碱基对间隔序列决定，该间隔序列分隔两个13碱基对的反向重复序列。位点特异性重组的结果或方向性由反应过程中两个同向位点的排列方式定义。间隔区内含有点突变、1碱基对插入或缺失的位点通常与未改变的位点以较低水平进行重组。相反，两个相同突变位点之间的重组（恢复了同源性）在所有情况下都能高效进行。含有10个或更多碱基对间隔序列的位点在所有条件下均无功能。一个重组位点，其中5个碱基对发生改变以产生完全对称的间隔序列，再次能高效重组，但仅与相同位点重组。此外，该反应产生多种新产物，这些产物只能由进行重组的两个位点的随机排列产生，即该反应不再表现出方向性。这些及其他结果表明，位点特异性重组的效率和方向性均取决于两个重组位点间隔序列之间的同源性。这进一步意味着，在该系统的位点特异性重组过程中，反应所涉及的两个位点间隔区之间关键的DNA - DNA相互作用在某个阶段会发生。只要保持同源性，特定的间隔序列本身似乎并不重要；因此，这些序列可能不参与FLP蛋白的识别。
