Infrared Spectroscopy Reveals the Preferred Motif Size and Local Disorder in Parallel Stranded DNA G‐Quadruplexes

Abstract

Infrared spectroscopy detects the formation of G-quadruplexes in guanine-rich nucleic acid sequences through shifts in the guanine C=O stretch mode. Here, we use ultrafast 2D infrared (IR) spectroscopy and isotope substitution to show that these shifts arise from vibrational delocalization among stacked Gquartets. This provides a direct measure of the sizes of locally ordered motifs in heterogeneous samples with substantial disordered regions. We find that parallel-stranded, potassiumbound DNA G-quadruplexes are limited to five consecutive G-quartets and 3-4 consecutive layers are preferred for longer polyguanine tracts. The resulting potassium-dependent Gquadruplex assembly landscape reflects the polyguanine tract lengths found in genomes, the ionic conditions prevalent in healthy mammalian cells, and the onset of structural disorder in disease states. Our study describes spectral markers that can be used to probe other G-quadruplex structures and provides insight into the fundamental limits of their formation in biological and artificial systems.

Probing the Hydrogen-Bonding Environment of Individual Bases in DNA Duplexes with Isotope-Edited Infrared Spectroscopy

“…A recent study used fully 15 N- and 13 C-labeled DNA bases to examine coupling in G-quadruplexes with ultrafast 2DIR spectroscopy. 41 …”

Membrane‐dependent amyloid aggregation of human BAX α9 (173–192)

“…Buffer backgrounds were subtracted and residual baseline correction was performed in MATLAB using low‐order polynomials. For 2D IR spectroscopy, ~17 μJ mid‐IR pulses with (~100 fs) were directed into a 2DQuick Array spectrometer (Phasetech Spectroscopy, Inc., Madison, WI) equipped with a 128 × 128 pixel MCT array detector as previously described 63 . All spectra were collected with perpendicular pump and probe polarizations and four‐frame phase cycling to reduce interference from pump scatter.…”

“…For 2D IR spectroscopy, 17 μJ mid-IR pulses with (100 fs) were directed into a 2DQuick Array spectrometer (Phasetech Spectroscopy, Inc., Madison, WI) equipped with a 128 × 128 pixel MCT array detector as previously described. 63 All spectra were collected with perpendicular pump and probe polarizations and four-frame phase cycling to reduce interference from pump scatter. Probe frequencies were calibrated to the absorbances of 4-nitrobenzaldehyde in dichloromethane (1,605; 1,676; and 1,709 cm −1 ), used as an external standard.…”

The Polarization Dependence of 2D IR Cross-Peaks Distinguishes Parallel-Stranded and Antiparallel-Stranded DNA G-quadruplexes

“…These arise from mixtures of signals from the constituent bases, and it is known from previous work that the ~1670 cm -1 feature contains contributions from guanine I modes, the ~1625 cm -1 feature contains contributions from thymine modes, and the ~1590 cm -1 feature contains contributions from guanine II modes. 27,29 was not certified by peer review) is the author/funder. All rights reserved.…”