関連する実験動画
Updated: May 11, 2026
07:37
Single-Molecule Real-Time Visualization of DNA Unwinding by CMG Helicase
Published on: September 27, 2024
RecBCDヘリケーゼを単一鎖DNAトランスロケーゼとして直接観察
1Department of Chemistry, National Taiwan University, Taiwan.
Journal of the American Chemical Society
|May 31, 2013
まとめ
DNA修復に不可欠なEscherichia coli RecBCD酵素は,単一鎖DNAに沿って両方向に移動することができます. この酵素は双方向転位を示しており,DNAの二重鎖破裂修復中に鎖を切り替えることができることを示唆しています.
科学分野:
- 分子生物学は分子生物学である.
- バイオケミストリー バイオケミストリー
- 遺伝学 遺伝学とは
背景:
- RecBCD酵素は,同類の再結合によってDNA二重鎖断裂 (DSB) 修復を開始する.
- 反対の極性を持つ2つのヘリコースモーターを備えています:RecB (3'-to-5') とRecD (5'-to-3').
研究 の 目的:
- 長い単一鎖DNA (ssDNA) 上のRecBCD酵素の転位能力を調査する.
- RecBCDが双方向的なssDNAトランスロカゼ活性を示し,複雑なDNA構造をナビゲートできるかどうかを判断する.
主な方法:
- 単一分子結合粒子運動 (TPM) 実験を活用した.
- ビーズラベルされた酵素を用いて,長いssDNA基板 (>200 nt) 上で視覚化されたRecBCD転位.
主要な成果:
- RecBCDは,3'-to-5'および5'-to-3'の両方向でssDNAに転位活性を示した.
- 酵素はssDNA領域に順調に転位し,ssDNA/dsDNA交差点で何らかの解離が観察されました.
- RecBCDは,相反の極性を持つssDNAの交互の領域をナビゲートすることができ,鎖交換能力を示しています.
結論:
- RecBCDは双方向的なssDNAトランスロカゼ活性を持っています.
- この酵素は,トランスロカゼの活動と,修復過程でDNA鎖をリタイヤする間の切り替えが可能である.
関連する概念動画
00:55DNA Helicases
DNA unwinding helicase enzymes are a type of motor protein. Motor proteins can translocate along filaments or polymers using energy generated from ATP hydrolysis. Helicases are involved in all the important cellular processes where DNA unwinding is required, such as DNA replication, repair, recombination, and transcription. They are present in all living organisms, but vary in their structure, function, and mechanism of action. For example, in prokaryotes, DnaB helicase binds and translocates...
02:31Homologous Recombination
The basic reaction of homologous recombination (HR) involves two chromatids that contain DNA sequences sharing a significant stretch of identity. One of these sequences uses a strand from another as a template to synthesize DNA in an enzyme-catalyzed reaction. The final product is a novel amalgamation of the two substrates. To ensure an accurate recombination of sequences, HR is restricted to the S and G2 phases of the cell cycle. At these stages, the DNA has been replicated already and the...
02:37Restarting Stalled Replication Forks
DNA replication is initiated at sites containing predefined DNA sequences known as origins of replication. DNA is unwound at these sites by the minichromosome maintenance (MCM) helicase and other factors such as Cdc45 and the associated GINS complex.The unwound single strands are protected by replication protein A (RPA) until DNA polymerase starts synthesizing DNA at the 5’ end of the strand in the same direction as the replication fork. To prevent the replication fork from falling apart, a...
01:02The DNA Replication Fork
An organism’s genome needs to be duplicated in an efficient and error-free manner for its growth and survival. The replication fork is a Y-shaped active region where two strands of DNA are separated and replicated continuously. The coupling of DNA unzipping and complementary strand synthesis is a characteristic feature of a replication fork. Organisms with small circular DNA, such as E. coli, often have a single origin of replication; therefore, they have only two replication forks, one in...