Taq DNA聚合酶功能区域的定位

Study on the Functional Domain of Taq DNA Polymerase

通过参Ｕ法定点突变产生了ＴａｑＤＮＡ聚合酶Ｎ端分别缺失３个，２３５个，２８７个和４４３个氨基酸的４个缺失体，利用Ｂａｌ－３１连续缺失法产生了ＴａｑＤＮＡ聚合酶的Ｃ端分别缺失了２个、１６个、２９个、３２个、３４个氨基酸的５个缺失体．经ＤＮＡ聚合酶活性测定表明Ｎ端缺失３个，２３５个，２８７个氨基酸后活力和完整的Ｔａｑ相近，而缺失４４３个氨基酸后则失去了ＤＮＡ聚合酶活力；Ｃ端的５个缺失体都失去了ＤＮＡ聚合酶活性．据此ＴａｑＤＮＡ聚合酶的功能区域被定位在２８７～８３２氨基酸之间．

Four truncators with deletion of 3,235,287 and 443 amino acids at N terminus of Taq DNA polymerase were obtained through ural mediated site direct mutagenesis.Five truncators with deletion of 2,16,29,32 and 34 amino acids at C terminus were obtained through continuous deletion with Bal 31 nuclease.Their DNA polymerase activity was measured.It was found that truncators with deletion of 3,235 and 287 amino acids at N terminus maintained the DNA polymerase activity and their specific activity was identical to that of native Taq DNA polymerase while other truncators with deletion at N terminus or C terminus lost their DNA polymerase activity.Based on these,the domain responsible for the DNA polymerase activity of Taq DNA polymerase was located between 287 amino acid and 832 amino acid.