Zaleski Piotr, Piekarowicz Andrzej
Institute of Microbiology, University of Warsaw, Miecznikowa 1, 02-096 Warsaw, Poland.
Microbiology (Reading). 2004 Nov;150(Pt 11):3773-3781. doi: 10.1099/mic.0.27225-0.
The gene encoding Dam methyltransferase of Haemophilus influenzae was mutagenized by the insertion of a chloramphenicol-resistance cassette into the middle of the Dam coding sequence. This mutant construct was introduced into the H. influenzae chromosome by transformation and selection for Cam(R) transformants. The authors have shown that several phenotypic properties, resistance to antibiotics, dyes and detergent as well as efficiency of transformation, depend on the Dam methylation state of the DNA. Although the major role of the methyl-directed mismatch repair (MMR) system is to repair postreplicative errors, it seems that in H. influenzae its effect is more apparent in repairing DNA damage caused by oxidative compounds. In the dam mutant treated with hydrogen peroxide, MMR is not targeted to newly replicated DNA strands and therefore mismatches are converted into single- and double-strand DNA breaks. This is shown by the increased peroxide sensitivity of the dam mutant and the finding that the sensitivity can be suppressed by a mutH mutation inactivating MMR. In the dam mutant treated with nitrofurazone the resulting damage is not converted into DNA breaks but the high sensitivity is also suppressed by a mutH mutation.
通过将氯霉素抗性盒插入流感嗜血杆菌Dam甲基转移酶编码基因的Dam编码序列中间，对该基因进行诱变。通过转化并选择氯霉素抗性（Cam(R)）转化体，将此突变构建体导入流感嗜血杆菌染色体。作者已表明，一些表型特性，如对抗生素、染料和去污剂的抗性以及转化效率，取决于DNA的Dam甲基化状态。尽管甲基导向错配修复（MMR）系统的主要作用是修复复制后错误，但在流感嗜血杆菌中，其作用似乎在修复由氧化化合物引起的DNA损伤方面更为明显。在用过氧化氢处理的dam突变体中，MMR不作用于新复制的DNA链，因此错配会转化为单链和双链DNA断裂。dam突变体对过氧化物的敏感性增加以及发现这种敏感性可被使MMR失活的mutH突变所抑制，都证明了这一点。在用呋喃西林处理的dam突变体中，产生的损伤不会转化为DNA断裂，但mutH突变也会抑制其高敏感性。
