New insight into the mechanism of DNA polymerase I revealed by single-molecule FRET studies of Klenow fragment

doi:10.1088/1674-1056/ac6ed5

中国物理B ›› 2022, Vol. 31 ›› Issue (8): 88701-088701.doi: 10.1088/1674-1056/ac6ed5

New insight into the mechanism of DNA polymerase I revealed by single-molecule FRET studies of Klenow fragment

Rokshana Parvin^1,2,†, Qi Jia(贾棋)^1,2,†, Jianbing Ma(马建兵)^1,2, Chunhua Xu(徐春华)^1,2, Ying Lu(陆颖)^1,2,‡, Fangfu Ye(叶方富)^1,2,§, and Ming Li(李明)^1,2,¶

1 Beijing National Laboratory for Condensed Matter Physics, Institute of Physics, Chinese Academy of Sciences, Beijing 100190, China;
2 School of Physical Sciences, University of Chinese Academy of Sciences, Beijing 100049, China

收稿日期:2022-04-18 修回日期:2022-05-07 接受日期:2022-05-12 出版日期:2022-07-18 发布日期:2022-08-02
通讯作者: Ying Lu, Fangfu Ye, Ming Li E-mail:yinglu@iphy.ac.cn;fye@iphy.ac.cn;mingli@iphy.ac.cn
基金资助:
Project supported by the National Natural Science Foundation of China (Grant No. 12090051), the CAS Key Research Program of Frontier Sciences (Grant Nos. QYZDJSSW-SYS014 and ZDBS-LY-SLH015), and the Youth Innovation Promotion Association of CAS (Grant No. 2017015).

New insight into the mechanism of DNA polymerase I revealed by single-molecule FRET studies of Klenow fragment

Rokshana Parvin^1,2,†, Qi Jia(贾棋)^1,2,†, Jianbing Ma(马建兵)^1,2, Chunhua Xu(徐春华)^1,2, Ying Lu(陆颖)^1,2,‡, Fangfu Ye(叶方富)^1,2,§, and Ming Li(李明)^1,2,¶

1 Beijing National Laboratory for Condensed Matter Physics, Institute of Physics, Chinese Academy of Sciences, Beijing 100190, China;
2 School of Physical Sciences, University of Chinese Academy of Sciences, Beijing 100049, China

Received:2022-04-18 Revised:2022-05-07 Accepted:2022-05-12 Online:2022-07-18 Published:2022-08-02
Contact: Ying Lu, Fangfu Ye, Ming Li E-mail:yinglu@iphy.ac.cn;fye@iphy.ac.cn;mingli@iphy.ac.cn
Supported by:
Project supported by the National Natural Science Foundation of China (Grant No. 12090051), the CAS Key Research Program of Frontier Sciences (Grant Nos. QYZDJSSW-SYS014 and ZDBS-LY-SLH015), and the Youth Innovation Promotion Association of CAS (Grant No. 2017015).

摘要/Abstract

摘要： We use single-molecule FRET and newly-developed D-loop techniques to investigate strand displacement activity of Klenow fragment (exo-) of DNA polymerase I in DNA sequences rich in guanine and cytosine (GC) bases. We find that there exist in the FRET traces numerous ascending jumps, which are induced by the backsliding of Klenow fragment on DNA chains. Our measurements show that the probability of backsliding is closely related to the GC-richness and dNTP concentration: increasing the GC-richness leads to an increase in the backsliding probability, and increasing the dNTP concentration however leads to a decrease in the backsliding probability. These results provide a new insight into the mechanism of DNA polymerase I.

关键词: smFRET, Klenow fragment, GC-richness, strand displacement

Abstract: We use single-molecule FRET and newly-developed D-loop techniques to investigate strand displacement activity of Klenow fragment (exo-) of DNA polymerase I in DNA sequences rich in guanine and cytosine (GC) bases. We find that there exist in the FRET traces numerous ascending jumps, which are induced by the backsliding of Klenow fragment on DNA chains. Our measurements show that the probability of backsliding is closely related to the GC-richness and dNTP concentration: increasing the GC-richness leads to an increase in the backsliding probability, and increasing the dNTP concentration however leads to a decrease in the backsliding probability. These results provide a new insight into the mechanism of DNA polymerase I.

Key words: smFRET, Klenow fragment, GC-richness, strand displacement

中图分类号: (Spectroscopic and microscopic techniques in biophysics and medical physics)

87.64.-t

87.80.Nj (Single-molecule techniques) 87.14.gk (DNA) 87.64.kv (Fluorescence)

Rokshana Parvin, Qi Jia(贾棋), Jianbing Ma(马建兵), Chunhua Xu(徐春华), Ying Lu(陆颖), Fangfu Ye(叶方富), and Ming Li(李明). New insight into the mechanism of DNA polymerase I revealed by single-molecule FRET studies of Klenow fragment[J]. 中国物理B, 2022, 31(8): 88701-088701.

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New insight into the mechanism of DNA polymerase I revealed by single-molecule FRET studies of Klenow fragment

New insight into the mechanism of DNA polymerase I revealed by single-molecule FRET studies of Klenow fragment

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