Use of UV spectroscopy for the study of nucleic acid cleavage by E. coli RNase H and restriction endonucleases.
A one-step spectrophotometric method for monitoring of nucleic acid cleavage by ribonuclease H from E. coli and type II restriction endonucleases has been proposed. It is based on recording of the increase in the UV absorbance at 260 nm during the course of enzymatic reaction. Duplexes stable under the reaction conditions were chosen as substrates for the enzymes being studied. In order to obtain duplex dissociation following their cleavage by the enzyme appreciate temperature conditions were selected. The spectrophotometric method may be applied for rapid testing of the nuclease activity in protein preparations as well as for precise quantitative analysis of nucleic acid degradation by enzymes. This method may be successfully employed in kinetic studies of nucleic acid-protein interactions.
Petrauskene
O V
OV
Department of Chemistry, Moscow State University, Russia.
Krynetskaya
N F
NF
Tashlitsky
V N
VN
Belkov
V M
VM
Kubareva
E A
EA
Gromova
E S
ES
Guschlbauer
W
W
Shabarova
Z A
ZA
D001483:Base Sequence; D004262:DNA Restriction Enzymes / Q000378:metabolism*; D004926:Escherichia coli / Q000201:enzymology*; D006868:Hydrolysis; D007700:Kinetics; D008969:Molecular Sequence Data; D009696:Nucleic Acids / Q000378:metabolism*; D015203:Reproducibility of Results; D016914:Ribonuclease H / Q000378:metabolism*; D013056:Spectrophotometry, Ultraviolet
D009696:Nucleic Acids; D004262:DNA Restriction Enzymes; D016914:Ribonuclease H