Biomaterial Database

A facile method for attaching nitroxide spin labels at the 5' terminus of nucleic acids. 2007

Gian Paola G Grant, and Peter Z Qin

Department of Chemistry, University of Southern California, LJS-251, 840 Downey Way, Los Angeles, CA 90089-0744, USA.

In site-directed spin labeling (SDSL), a nitroxide moiety containing a stable, unpaired electron is covalently attached to a specific site within a macromolecule, and structural and dynamic information at the labeling site is obtained via electron paramagnetic resonance (EPR) spectroscopy. Successful SDSL requires efficient site-specific incorporation of nitroxides. Work reported here presents a new method for facile nitroxide labeling at the 5' terminus of nucleic acids of arbitrary sizes. T4-polynucleotide kinase was used to enzymatically substitute a phosphorothioate group at the 5' terminus of a nucleic acid, and the resulting phosphorothioate was then reacted with an iodomethyl derivative of a nitroxide. The method was successfully demonstrated on both chemically synthesized and naturally occurring nucleic acids. The attached nitroxides reported duplex formation as well as tertiary folding of nucleic acids, indicating that they serve as a valid probe in nucleic acid studies.

UI	MeSH Term	Description	Entries
D009589	Nitrogen Oxides	Inorganic oxides that contain nitrogen.	Nitrogen Oxide,Oxide, Nitrogen,Oxides, Nitrogen
D011116	Polynucleotide 5'-Hydroxyl-Kinase	An enzyme that catalyzes the transfer of a phosphate group to the 5'-terminal hydroxyl groups of DNA and RNA. EC 2.7.1.78.	Polynucleotide Hydroxylkinase,Polynucleotide Kinase,5'-Hydroxylpolynucleotide Kinase,DNA 5'-Hydroxylkinase,DNA Kinase,Polynucleotide 5'-Hydroxyl Kinase,Polynucleotide Hydroxykinase,5' Hydroxylpolynucleotide Kinase,5'-Hydroxyl Kinase, Polynucleotide,5'-Hydroxyl-Kinase, Polynucleotide,5'-Hydroxylkinase, DNA,DNA 5' Hydroxylkinase,Hydroxykinase, Polynucleotide,Hydroxylkinase, Polynucleotide,Kinase, 5'-Hydroxylpolynucleotide,Kinase, DNA,Kinase, Polynucleotide,Kinase, Polynucleotide 5'-Hydroxyl,Polynucleotide 5' Hydroxyl Kinase
D004247	DNA	A deoxyribonucleotide polymer that is the primary genetic material of all cells. Eukaryotic and prokaryotic organisms normally contain DNA in a double-stranded state, yet several important biological processes transiently involve single-stranded regions. DNA, which consists of a polysugar-phosphate backbone possessing projections of purines (adenine and guanine) and pyrimidines (thymine and cytosine), forms a double helix that is held together by hydrogen bonds between these purines and pyrimidines (adenine to thymine and guanine to cytosine).	DNA, Double-Stranded,Deoxyribonucleic Acid,ds-DNA,DNA, Double Stranded,Double-Stranded DNA,ds DNA
D004578	Electron Spin Resonance Spectroscopy	A technique applicable to the wide variety of substances which exhibit paramagnetism because of the magnetic moments of unpaired electrons. The spectra are useful for detection and identification, for determination of electron structure, for study of interactions between molecules, and for measurement of nuclear spins and moments. (From McGraw-Hill Encyclopedia of Science and Technology, 7th edition) Electron nuclear double resonance (ENDOR) spectroscopy is a variant of the technique which can give enhanced resolution. Electron spin resonance analysis can now be used in vivo, including imaging applications such as MAGNETIC RESONANCE IMAGING.	ENDOR,Electron Nuclear Double Resonance,Electron Paramagnetic Resonance,Paramagnetic Resonance,Electron Spin Resonance,Paramagnetic Resonance, Electron,Resonance, Electron Paramagnetic,Resonance, Electron Spin,Resonance, Paramagnetic
D001671	Biochemistry	The study of the composition, chemical structures, and chemical reactions of living things.
D012313	RNA	A polynucleotide consisting essentially of chains with a repeating backbone of phosphate and ribose units to which nitrogenous bases are attached. RNA is unique among biological macromolecules in that it can encode genetic information, serve as an abundant structural component of cells, and also possesses catalytic activity. (Rieger et al., Glossary of Genetics: Classical and Molecular, 5th ed)	RNA, Non-Polyadenylated,Ribonucleic Acid,Gene Products, RNA,Non-Polyadenylated RNA,Acid, Ribonucleic,Non Polyadenylated RNA,RNA Gene Products,RNA, Non Polyadenylated
D012360	RNA, Transfer, Phe	A transfer RNA which is specific for carrying phenylalanine to sites on the ribosomes in preparation for protein synthesis.	Phenylalanine-Specific tRNA,Transfer RNA, Phe,tRNAPhe,tRNA(Phe),Phe Transfer RNA,Phenylalanine Specific tRNA,RNA, Phe Transfer,tRNA, Phenylalanine-Specific
D013113	Spin Labels	Molecules which contain an atom or a group of atoms exhibiting an unpaired electron spin that can be detected by electron spin resonance spectroscopy and can be bonded to another molecule. (McGraw-Hill Dictionary of Chemical and Technical Terms, 4th ed)	Spin Label,Label, Spin,Labels, Spin
D013873	Thionucleotides	Nucleotides in which the base moiety is substituted with one or more sulfur atoms.