biology Review Volumetric Properties of Four-Stranded DNA Structures Tigran V. Chalikian * and Robert B. Macgregor, Jr.   Citation: Chalikian, T.V.; Macgregor, R.B., Jr. Volumetric Properties of Four-Stranded DNA Structures. Biology 2021, 10, 813. https:// doi.org/10.3390/biology10080813 Academic Editor: Dmitri Davydov Received: 15 July 2021 Accepted: 19 August 2021 Published: 22 August 2021 Publisher’s Note: MDPI stays neutral with regard to jurisdictional claims in published maps and institutional affil- iations. Copyright: © 2021 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https:// creativecommons.org/licenses/by/ 4.0/). Department of Pharmaceutical Sciences, Leslie Dan Faculty of Pharmacy, University of Toronto, 144 College Street, Toronto, ON M5S 3M2, Canada; rob.macgregor@utoronto.ca * Correspondence: t.chalikian@utoronto.ca; Tel.: +1-416-946-3715; Fax: +1-416-978-8511 Simple Summary: The volumetric properties of biomolecules define their pressure stability, while also characterizing their intrinsic and hydration properties. In this paper, we review the recent progress in volumetric investigations of G-quadruplexes and i-motifs, four-stranded secondary structures of DNA that have been found in the cell and implicated in regulatory genomic functions. Although the volumetric studies of G-quadruplexes and i-motifs are still in their nascent state, the data on volume, expansibility, and compressibility accumulated to date have begun to provide insights into the balance of forces governing the stability of these non-canonical structures. We present the available volumetric data and discuss how they can be rationalized in terms of intra-and intermolecular interactions involving G-quadruplexes and i-motifs including their solute-solvent interactions. Abstract: Four-stranded non-canonical DNA structures including G-quadruplexes and i-motifs have been found in the genome and are thought to be involved in regulation of biological function. These structures have been implicated in telomere biology, genomic instability, and regulation of transcription and translation events. To gain an understanding of the molecular determinants underlying the biological role of four-stranded DNA structures, their biophysical properties have been extensively studied. The limited libraries on volume, expansibility, and compressibility accumulated to date have begun to provide insights into the molecular origins of helix-to-coil and helix-to-helix conformational transitions involving four-stranded DNA structures. In this article, we review the recent progress in volumetric investigations of G-quadruplexes and i-motifs, emphasizing how such data can be used to characterize intra-and intermolecular interactions, including solvation. We describe how volumetric data can be interpreted at the molecular level to yield a better understanding of the role that solute–solvent interactions play in modulating the stability and recognition events of nucleic acids. Taken together, volumetric studies facilitate unveiling the molecular determinants of biological events involving biopolymers, including G-quadruplexes and i-motifs, by providing one more piece to the thermodynamic puzzle describing the energetics of cellular processes in vitro and, by extension, in vivo. Keywords: G-quadruplex; i-motif; volumetric properties; pressure-temperature phase diagram; thermodynamics 1. Introduction DNA molecules rich in guanine are prone to folding into four-stranded G-quadruplex structures, while cytosine-rich molecules tend to fold into four-stranded i-motif structures at slightly acidic pH [1–9]. G-quadruplexes are formed by stacking of two or more G- tetrads on top of each other. A G-tetrad represents a cyclic planar construct in which four guanine bases are linked together via Hoogsteen hydrogen bonds as shown in Figure 1a. The stacking results in the formation of a central cavity in which mono- or divalent cations are coordinated to the O6 atoms of guanines [1–3,7,10–12]. Sodium and potassium are the two biologically most relevant cations stabilizing G-quadruplex structures. The four consecutive G-runs involved in the formation of stacked tetrads in an intramolecular G- quadruplex are connected to each other via three single stranded linkers known as loops. Biology 2021, 10, 813. https://doi.org/10.3390/biology10080813 https://www.mdpi.com/journal/biology