Determination of base pairing in ribonucleic acid by Fourier-transform infrared spectrometry: yeast ribosomal 5S ribonucleic acid.
Fourier-transform infrared (FT-IR) spectra of yeast ribosomal 5S RNA have been acquired at several temperatures between 30 and 90 degrees C. The difference spectrum between 90 (bases unstacked) and 30 degrees C (bases stacked) provides a measure of base stacking in the RNA. Calibration difference spectra corresponding to stacking of G-C or A-U pairs are obtained from "reference" FT-IR spectra of poly(rG) X poly(rC) minus 5'-GMP and 5'-CMP or poly(rA) X poly(rU) minus 5'-AMP and 5'-UMP. The best fit linear combination of the calibration G-C and A-U difference spectra to the 5S RNA (90-30 degrees C) difference spectrum leads to a total of 25 +/- 3 base pairs (17 G-C pairs + 8 A-U pairs) for the native yeast 5S RNA in the absence of Mg2+. In the presence of Mg2+, an additional six base pairs are detected by FT-IR (one G-C and five A-U). FT-IR melting curve midpoints show that A-U and G-C pairs melt together (65 and 63 degrees C) in the presence of Mg2+ but A-U pairs melt before G-C pairs (47 vs. 54 degrees C) in the absence of Mg2+.
D001482:Base Composition; D001483:Base Sequence; D005583:Fourier Analysis; D008274:Magnesium; D015636:Magnesium Chloride; D009690:Nucleic Acid Conformation; D012331:RNA, Fungal / Q000302:isolation & purification; D012335:RNA, Ribosomal / Q000302:isolation & purification; D012441:Saccharomyces cerevisiae; D013055:Spectrophotometry, Infrared
D012331:RNA, Fungal; D012335:RNA, Ribosomal; D015636:Magnesium Chloride; D008274:Magnesium