The Denaturation Transition of DNA in Mixed Solvents

INTRODUCTION

DNA is a biopolymer formed of backbone phosphates, linked to desoxyribose sugars and side group amine bases (1). The charged phosphate groups are hydrophilic, the desoxyribose sugar groups are mostly hydrophobic, and the amine bases contain both hydrophobic and hydrophilic groups. This is based on water binding ability as referenced below. DNA forms a helical structure which is stable because of the stacking of the amine bases and of the hydrogen bonding between them. The helix phase melts into disordered coils under various conditions including temperature increase.

Most research on DNA has been in aqueous media at low concentration (2-10). Crystallouraphic measurements give precise sizes for the DNA helical structure: 3.4 [Angstrom] average distance between adjacent phosphates, 34 [Angstrom] for the helix pitch, and 20 [Angstrom] for the helix diameter (3). The ultraviolet (UV) absorption spectroscopy method (3,4) has proven valuable for the characterization of the helix-to-coil transition. Some investigations used non-aqueous solvents such as ethylene glycol (6,9). Other analytical methods such as NMR or infrared (IR) spectroscopy (8,10) have been used to estimate hydration properties. It was found for instance that a specific number of water molecules are bound to each DNA nucleotide (10) (5 molecules per amine basepair, 4 molecules per phosphate, and 2 molecules pertlesoxyrihose sugar group). X-ray and neutron fiber diffraction yielded information about water hydration as well (11). In Fuller et al. (11), it is stated "challenges in developing alternatives to a water environment can he expected to be very severe". This article reports a study of the ethylene glycol alternative.

The helix-to-coil melting transition in DNA has been the subject of a large number of investigations in the literature, among which are review articles (12-14). Cheng and Pettitt (13) contains a selected literature review of experimental and modeling efforts. Stability of the helix structure is governed by the amine base stacking and the base-pairing through hydrogen-bonding. Factors like temperature. DNA concentration, pH, salt concentration and solvent mixtures affect the helix-to-coil transition (13). The helix-to-coil transition occurs in transcription and replication of DNA. and is also is a key aspect of the polymerase chain reaction in biotechnology.

METHODS AND MATERIALS

The focus of our investigations is on the characterization ot the helix-to-coil dcnaturation transition of DNA (1) in cthylcnc glycol and (2) in water/ ethylene glycol mixtures. Two measurement melhixls are used: UV light absorption spectroscopy to characterize the helix-to-coil melting temperalure and small-angle neutron scattering (SANS) to monitor structural changes across the melting transition.