HpaI 限制-修饰基因的克隆与表达

Ito H, Shimato H, Sadaoka A, Kotani H, Kimizuka F, Kato I

Bioproducts Development Center, Takara Shuzo Co., Ltd, Shiga, Japan.

Nucleic Acids Res. 1992 Feb 25;20(4):705-9. doi: 10.1093/nar/20.4.705.

The genes from Haemophilus parainfluenzae encoding the HpaI restriction-modification system were cloned and expressed in Escherichia coli. From the DNA sequence, we predicted the HpaI endonuclease (R.HpaI) to have 254 amino acid residues (Mr 29,630) and the HpaI methyltransferase (M.HpaI) to have 314 amino acid residues (37,390). The R.HpaI and M.HpaI genes overlapped by 16 base pairs on the chromosomal DNA. The genes had the same orientation. The clone, named E. coli HB101-HPA2, overproduced R.HpaI. R.HpaI activity from the clone was 100-fold that from H. parainfluenzae. The amino acid sequence of M.HpaI was compared with those of other type II methyltransferases.

编码副流感嗜血杆菌HpaI限制修饰系统的基因被克隆并在大肠杆菌中表达。根据DNA序列，我们预测HpaI核酸内切酶（R.HpaI）有254个氨基酸残基（分子量29,630），HpaI甲基转移酶（M.HpaI）有314个氨基酸残基（37,390）。R.HpaI和M.HpaI基因在染色体DNA上重叠16个碱基对。这些基因具有相同的方向。该克隆名为大肠杆菌HB101-HPA2，过量产生R.HpaI。该克隆的R.HpaI活性是副流感嗜血杆菌的100倍。将M.HpaI的氨基酸序列与其他II型甲基转移酶的序列进行了比较。