Kumar N V, Varshney U
Centre for Genetic Engineering, Indian Institute of Science, Bangalore.
Nucleic Acids Res. 1994 Sep 11;22(18):3737-41. doi: 10.1093/nar/22.18.3737.
Kinetic parameters for uracil DNA glycosylase (E. coli)-catalysed excision of uracil from DNA oligomers containing dUMP in different structural contexts were determined. Our results show that single-stranded oligonucleotides (unstructured) are used as somewhat better substrates than the double-stranded oligonucleotides. This is mainly because of the favourable Vmax value of the enzyme for single-stranded substrates. More interestingly, however, we found that uracil release from loop regions of DNA hairpins is extremely inefficient. The poor efficiency with which uracil is excised from loop regions is a result of both increased Km and lowered Vmax values. This observation may have significant implications in uracil DNA glycosylase-directed repair of DNA segments that can be extruded as hairpins. In addition, these studies are useful in designing oligonucleotides for various applications in DNA research where the use of uracil DNA glycosylase is sought.
测定了尿嘧啶DNA糖基化酶（大肠杆菌）催化从不同结构背景下含有dUMP的DNA寡聚物中切除尿嘧啶的动力学参数。我们的结果表明，单链寡核苷酸（无结构）用作底物时比双链寡核苷酸稍好。这主要是因为该酶对单链底物具有有利的Vmax值。然而，更有趣的是，我们发现从DNA发夹环区域释放尿嘧啶的效率极低。从环区域切除尿嘧啶的效率低下是Km增加和Vmax值降低的结果。这一观察结果可能对尿嘧啶DNA糖基化酶指导的可挤出成发夹的DNA片段修复具有重要意义。此外，这些研究对于设计用于寻求使用尿嘧啶DNA糖基化酶的DNA研究中各种应用的寡核苷酸很有用。
