Adams C W, Nanassy O, Johnson R C, Hughes K T
Department of Microbiology, University of Washington, Seattle 98195, USA.
Mol Microbiol. 1997 Jun;24(6):1235-47. doi: 10.1046/j.1365-2958.1997.4141789.x.
The Hin recombinase mediates the site-specific inversion of a segment of the Salmonella chromosome between two flanking 26bp hix DNA recombination sites. Mutations in two amino acid residues, R43 and R69 of the catalytic domain of the Hin recombinase, were identified that can compensate for loss of binding resulting from elimination of certain major and minor groove contacts within the hix recombination sites. With one exception, the R43 and R69 mutants were also able to bind a hix sequence with an additional 4bp added to the centre of the site, unlike wild-type Hin. Purified Hin mutants R43H and R69C had both partial cleavage and inversion activities in vitro while mutants R43L, R43C, R69S, and R69P had no detectable cleavage and inversion activities. These data support a model in which the catalytic domain plays a role in DNA-binding specificity, and suggest that the arginine residues at positions 43 and 69 function to position the Hin recombinase on the DNA for a step in the recombination reaction which occurs either at and/or prior to DNA cleavage.
Hin 重组酶介导沙门氏菌染色体一段序列在两个侧翼 26bp 的 hix DNA 重组位点之间进行位点特异性倒位。已鉴定出 Hin 重组酶催化结构域中两个氨基酸残基（R43 和 R69）发生的突变，这些突变能够补偿因 hix 重组位点内某些主要和次要沟接触消除而导致的结合丧失。除了一个例外，与野生型 Hin 不同，R43 和 R69 突变体还能够结合在位点中心额外添加了 4bp 的 hix 序列。纯化的 Hin 突变体 R43H 和 R69C 在体外具有部分切割和倒位活性，而突变体 R43L、R43C、R69S 和 R69P 没有可检测到的切割和倒位活性。这些数据支持了一种模型，即催化结构域在 DNA 结合特异性中起作用，并表明 43 位和 69 位的精氨酸残基在重组反应中 DNA 切割时和/或之前的步骤中，发挥将 Hin 重组酶定位在 DNA 上的功能。
