Determination of DNA structure in solution: enzymatic deuteration of the ribose 2' carbon.
An enzymatic solution to the problem of obtaining 13C/15N-labeled nucleotides that are deuterated uniquely at the H2' ' position within the ribose ring is presented. Selective deuteration occurs with an overall yield of >80%. The deuteron at the H2' ' position allows measurement of the scalar and residual dipolar couplings for the bond vectors attached to the C2' carbon of each ribose sugar. These data allow the accurate determination of sugar conformation. Interesting DNA double helices of 2-3 turns are now within the reach of solution NMR spectroscopy. As an example, these labeled nucleotides are incorporated uniquely at positions 6-14 in a 20-bp DNA sequence containing the adenovirus major late promoter.
MacDonald
Douglas
D
Department of Chemistry, University of Pennsylvania, Philadelphia, Pennsylvania 19104-6323, USA.
Lu
Ponzy
P
D002247:Carbon Isotopes; D004247:DNA / Q000737:chemistry*; D003854:Deoxyribonucleotides / Q000737:chemistry*; D003856:Deoxyuracil Nucleotides / Q000737:chemistry; D003903:Deuterium / Q000737:chemistry*; D004926:Escherichia coli / Q000201:enzymology; D007553:Isotope Labeling / Q000379:methods; D007778:Lactobacillus / Q000201:enzymology; D009587:Nitrogen Isotopes; D019906:Nuclear Magnetic Resonance, Biomolecular / Q000379:methods; D009690:Nucleic Acid Conformation; D011755:Pyrophosphatases / Q000737:chemistry / Q000378:metabolism; D012264:Ribonucleotide Reductases / Q000737:chemistry / Q000378:metabolism; D012266:Ribose / Q000737:chemistry*
D002247:Carbon Isotopes; D003854:Deoxyribonucleotides; D003856:Deoxyuracil Nucleotides; D009587:Nitrogen Isotopes; C027078:deoxyuridine triphosphate; D012266:Ribose; D004247:DNA; D003903:Deuterium; D012264:Ribonucleotide Reductases; C057295:ribonucleoside-triphosphate reductase; D011755:Pyrophosphatases; C020020:dUTP pyrophosphatase