Gunn J S, Stein D C
University of Maryland, Department of Microbiology, College Park, MD 20142, USA.
Nucleic Acids Res. 1997 Oct 15;25(20):4147-52. doi: 10.1093/nar/25.20.4147.
Strains of Neisseria gonorrhoeae possess numerous restriction-modification (R-M) systems. One of these systems, which has been found in all strains tested, encodes the S. NgoVIII specificity (5'TCACC 3') R-M system. We cloned two adjacent methyltransferase genes (dcmH and damH), each encoding proteins whose actions protect DNA from digestion by R.HphI or R.Ngo BI (5'TCACC 3'). The damH gene product is a N 6-methyladenine methyltransferase that recognizes this sequence. We constructed a plasmid containing multiple copies of the S.NgoVIII sequence, grew it in the presence of damH and used the HPLC to demonstrate the presence of N 6-methyladenine in the DNA. A second plasmid, containing overlapping damH and Escherichia coli dam recognition sequences in combination with various restriction digests, was used to identify which adenine in the recognition sequence was modified by damH. The predicted dcmH gene product is homologous to 5-methylcytosine methyltransferases. The products of both the dcmH and damH genes, as well as an open reading frame downstream of the damH gene are highly similar to the Haemophilus parahaemolyticus hphIMC , hphIMA and hphIR gene products, encoding the Hph I Type IIs R-M system. The S.NgoVIII R-M genes are flanked by a 97 bp direct repeat that may be involved in the mobility of this R-M system.
淋病奈瑟菌菌株拥有众多限制修饰(R-M)系统。在所有测试菌株中均发现的其中一个系统,编码S. NgoVIII特异性(5'TCACC 3')R-M系统。我们克隆了两个相邻的甲基转移酶基因(dcmH和damH),每个基因编码的蛋白质可保护DNA不被R.HphI或R.Ngo BI(5'TCACC 3')消化。damH基因产物是一种识别该序列的N6-甲基腺嘌呤甲基转移酶。我们构建了一个含有多个S.NgoVIII序列拷贝的质粒,在damH存在的情况下培养它,并使用高效液相色谱法证明DNA中存在N6-甲基腺嘌呤。第二个质粒含有重叠的damH和大肠杆菌dam识别序列,并结合各种限制性酶切,用于鉴定识别序列中的哪个腺嘌呤被damH修饰。预测的dcmH基因产物与5-甲基胞嘧啶甲基转移酶同源。dcmH和damH基因的产物,以及damH基因下游的一个开放阅读框,与副溶血嗜血杆菌hphIMC、hphIMA和hphIR基因产物高度相似,后者编码Hph I I型R-M系统。S.NgoVIII R-M基因两侧有一个97 bp的直接重复序列,可能参与该R-M系统的移动。