Regulation of DNA nucleases by molecular crowding.
Here, we examined the effects of molecular crowding on the function, structure and stability of nucleases. We found that the hydrolysis of a 29-mer double-stranded DNA by the endonucleases DNase I and S1 nuclease was substantially enhanced by molecular crowding using polyethylene glycol (PEG); however, molecular crowding had little effect on hydrolysis by exo III and exo I exonucleases. Moreover, kinetic analysis showed that the maximum velocity for the reaction of DNase I at 25 degrees C was increased from 0.1 to 2.7 microM/min by molecular crowding with 20% (w/v) PEG, whereas that of exonuclease I at 37 degrees C decreased from 2.2 to 0.4 microM/min. In contrast, molecular crowding did not significantly affect the Michaelis constant of DNase I or exonuclease I. These results indicate that molecular crowding has different effects on the catalytic activities of exonucleases and endonucleases.
Sasaki
Yoshiharu
Y
Frontier Institute for Biomolecular Engineering Research (FIBER), Konan University, 8-9-1 Okamoto, Higashinada-ku, Kobe 658-8501, Fine Co., Ltd., 5-7-8 Shimoshinjo, Higashiyodogawa-ku, Osaka.
Miyoshi
Daisuke
D
Sugimoto
Naoki
N
D004247:DNA / Q000378:metabolism; D003850:Deoxyribonuclease I / Q000378:metabolism; D004706:Endodeoxyribonucleases / Q000378:metabolism*; D005090:Exodeoxyribonucleases / Q000378:metabolism*; D006868:Hydrolysis; D007700:Kinetics; D011092:Polyethylene Glycols / Q000737:chemistry; D015719:Single-Strand Specific DNA and RNA Endonucleases / Q000378:metabolism; D013816:Thermodynamics
D011092:Polyethylene Glycols; D004247:DNA; D004706:Endodeoxyribonucleases; D005090:Exodeoxyribonucleases; C028450:exodeoxyribonuclease I; C020133:exodeoxyribonuclease III; D003850:Deoxyribonuclease I; D015719:Single-Strand Specific DNA and RNA Endonucleases