Anton B P, Heiter D F, Benner J S, Hess E J, Greenough L, Moran L S, Slatko B E, Brooks J E
New England Biolabs, Beverly, MA 01915, USA.
Gene. 1997 Mar 10;187(1):19-27. doi: 10.1016/s0378-1119(96)00638-5.
Bg/II, a type II restriction-modification (R-M) system from Bacillus globigii, recognizes the sequence 5'-AGATCT-3'. The system has been cloned into E. coli in multiple steps: first the methyltransferase (MTase) gene, bglIIM, was cloned from B. globigii RUB561, a variant containing an inactivated endonuclease (ENase) gene (bglIIR). Next the ENase protein (R.BglII) was purified to homogeneity from RUB562, a strain expressing the complete R-M system. Oligonucleotide probes specific for the 5' end of the gene were then synthesized and used to locate bglIIR, and the gene was isolated and cloned in a subsequent step. The nucleotide sequence of the system has been determined, and several interesting features have been found. The genes are tandemly arranged, with bglIIR preceding bglIIM. The amino acid sequence of M.BglII is compared to those of other known MTases. A third gene encoding a protein with sequence similarity to known C elements of other R-M systems is found upstream of bglIIR. This is the first instance of a C gene being associated with an R-M system where the R and M genes are collinear. In addition, open reading frames (ORFs) resembling genes involved with DNA mobility are found in close association with BglII. These may shed light on the evolution of the R-M system.
Bg/II是一种来自球状芽孢杆菌的II型限制-修饰（R-M）系统，识别序列5'-AGATCT-3'。该系统已通过多个步骤克隆到大肠杆菌中：首先，从球状芽孢杆菌RUB561中克隆甲基转移酶（MTase）基因bglIIM，RUB561是一种含有失活内切核酸酶（ENase）基因（bglIIR）的变体。接下来，从表达完整R-M系统的菌株RUB562中纯化出均一的ENase蛋白（R.BglII）。然后合成了对该基因5'端特异的寡核苷酸探针，并用于定位bglIIR，随后分离并克隆了该基因。已确定了该系统的核苷酸序列，并发现了几个有趣的特征。这些基因串联排列，bglIIR在bglIIM之前。将M.BglII的氨基酸序列与其他已知MTase的氨基酸序列进行了比较。在bglIIR上游发现了第三个基因，其编码的蛋白质与其他R-M系统的已知C元件具有序列相似性。这是C基因与R和M基因共线的R-M系统相关联的首例。此外，发现与DNA移动性相关的类似基因的开放阅读框（ORF）与BglII紧密相关。这些可能有助于揭示R-M系统的进化。
