Direct observation of positive supercoils introduced by reverse gyrase through atomic force microscopy |
| |
Authors: | Dawei Li Bei Lv Qiang wang Yun Liu Qiang Zhuge |
| |
Institution: | 1. Key Lab of Forest Genetics and Biotechnology, Nanjing Forestry University, 159 Longpan Road, Nanjing 210037, China;2. Jiangsu Key Laboratory for Biofunctional Molecules, College of Life Science and Chemistry, Jiangsu Second Normal University, Nanjing 210037, China |
| |
Abstract: | Reverse gyrase is a hyperthermophilic enzyme that can introduce positive supercoiling in substrate DNA. It is showed in our studies that positive DNA supercoils were induced in both pBR322 vector and an artificially synthesized mini-plasmid DNA by reverse gyrase. The left-handed structures adopted by positively supercoiled DNA molecules could be identified from their right-handed topoisomers through atomic force microscopic examination. Additional structural comparisons revealed that positively supercoiled DNA molecule AFM images exhibited increased contour lengths. Moreover, enzymatic assays showed that the positively supercoiled DNA could not be cleaved by T7 endonuclease. Together, this suggests that the overwound structure of positive supercoils could prevent genomic duplex DNA from randomly forming single-stranded DNA regions and intra-stranded secondary structures. |
| |
Keywords: | Positive supercoil Reverse gyrases DNA writhe AFM examination |
本文献已被 ScienceDirect 等数据库收录! |