Fast-scanning atomic force microscopy reveals the molecular mechanism of DNA cleavage by ApaI endonuclease.
Newly developed fast-scanning atomic force microscopy (AFM) allows the dissection of molecular events such as DNA-enzyme reactions at the single-molecule level. With this novel technology, a model is proposed of the DNA cleavage reaction by a type IIP restriction endonuclease ApaI. Detailed analyses revealed that ApaI bound to DNA as a dimer and slid along DNA in a one-dimensional diffusion manner. When it encountered a specific DNA sequence, the enzyme halted for a moment to digest the DNA. Immediately after digestion, the ApaI dimer separated into two monomers, each of which remained on the DNA end and then dissociated from the DNA end. Thus, fast-scanning AFM is a powerful tool to aid the understanding of protein structures and dynamics in biological reactions at the single-molecule level in sub-seconds.
Yokokawa
M
M
Laboratory of Plasma Membrane & Nuclear Signaling, Kyoto University Graduate School of Biostudies, Kitashirakawa Oiwake-cho, Sakyo-ku, Kyoto 606-8502, Japan. yokokawa@lif.kyoto-u.ac.jp
Yoshimura
S H
SH
Naito
Y
Y
Ando
T
T
Yagi
A
A
Sakai
N
N
Takeyasu
K
K
D001665:Binding Sites; D003198:Computer Simulation; D003199:Computer Systems; D004247:DNA / Q000737:chemistry* / Q000648:ultrastructure*; D004268:DNA-Binding Proteins / Q000737:chemistry / Q000648:ultrastructure; D015252:Deoxyribonucleases, Type II Site-Specific / Q000737:chemistry / Q000378:metabolism* / Q000648:ultrastructure*; D004789:Enzyme Activation; D018625:Microscopy, Atomic Force / Q000379:methods*; D008956:Models, Chemical; D008958:Models, Molecular; D009038:Motion; D009690:Nucleic Acid Conformation; D011485:Protein Binding; D011487:Protein Conformation
D004268:DNA-Binding Proteins; D004247:DNA; D015252:Deoxyribonucleases, Type II Site-Specific; C038485:GGGCCC-specific type II deoxyribonucleases