人乳铁蛋白阳离子多肽重组Taq DNA聚合酶的研究

Study on Modification of Taq DNA Polymerase by Replacing of its N-terminal 289 Amino Acids with Cationic Peptide of Human Lactoferrin

摘要:

Taq DNA聚合酶的模板DNA结合与DNA合成催化功能分别位于酶蛋白的不同区域，运用基因操作技术对某些区域进行替换重组是创建新型Taq酶的途径。人乳铁蛋白N-端阳离子多肽（HL-N）具较强的DNA结合能力,已被广泛研究和报道。本文研究了用人乳铁蛋白阳离子多肽构建嵌合重组Taq酶，和重组酶对DNA合成加工、血清耐受性等性质的影响。数据显示，HL-N重组的Taq酶在DNA扩增速度、扩增物产量、以及对反应体系中血清浓度的耐受性都得到明显提高。

Abstract:

Taq DNA polymerase consists of isolated functional domains responsible for binding of template DNA and catalysis of DNA synthesis, respectively. Previous research showed that the DNA-binding domain can be replaced by other protein or motif with higher DNA-binding affinity, resulting a modification in several properties of the recombinant polymerase．In the present study, a recombinant Taq DNA polymerase was constructed by replacing the N-terminal 289 amino acids of Taq DNA polymerase with cationic peptide of human lactoferrin (NL-N)．Some enzymatic properties of the chimerical recombinant DNA polymerase, such as the processivity, 5'-3' exonuclease activity, and tolerance to serum concentration in PCR reaction system, were characterized experimentally and the testing data showed that the modification of DNA-binding domain with NL-N improved the polymerase in several aspects.