DNA–Gelatin Complex Coacervation, UCST and First-Order Phase Transition of Coacervate to Anisotropic ion gel in 1-Methyl-3-octylimidazolium Chloride Ionic Liquid Solutions

Abstract

Study of kinetics of complex coacervation occurring in aqueous 1-octyl-3-methylimidazolium chloride ionic liquid solution of low charge density polypeptide (gelatin A) and 200 base pair DNA, and thermally activated coacervate into anisotropic gel transition, is reported here. Associative interaction between DNA and gelatin A (GA) having charge ratio (DNA:GA = 16:1) and persistence length ratio (5:1) was studied at fixed DNA (0.005% (w/v)) and varying GA concentration (C(GA) = 0-0.25% (w/v)). The interaction profile was found to be strongly hierarchical and revealed three distinct binding regions: (i) Region I showed DNA-condensation (primary binding) for C(GA) < 0.10% (w/v), the DNA ζ potential decrease from -80 to -5 mV (95%) (partial charge neutralization), and a size decrease by ≈60%. (ii) Region II (0.10 < C(GA) < 0.15% (w/v)) indicated secondary binding, a 4-fold turbidity increase, a ζ potential decrease from -5 to 0 mV (complete charge neutralization), which resulted in the appearance of soluble complexes and initiation of coacervation. (iii) Region III (0.15 < C(GA) < 0.25% (w/v)) revealed growth of insoluble complexes followed by precipitation. The hydration of coacervate was found to be protein concentration specific in Raman studies. The binding profile of DNA-GA complex with IL concentration revealed optimum IL concentration (=0.05% (w/v)) was required to maximize the interactions. Small angle neutron scattering (SANS) data of coacervates gave static structure factor profiles, I(q) versus wave vector q, that were remarkably similar and invariant of protein concentration. This data could be split into two distinct regions: (i) for 0.0173 < q < 0.0353 Å(-1), I(q) ~ q(-α) with α = 1.35-1.67, and (ii) for 0.0353 < q < 0.35 Å(-1), I(q) = I(0)/(1 + q(2)ξ(2)). The correlation length found was ξ = 2 ± 0.1 nm independent of protein concentration. The viscoelastic length (≈8 nm) was found to have value close to the persistence length of the protein (≈10 nm). Rheology data indicated that the coacervate phase resided close to the gelation state of the protein. Thus, on a heating-cooling cycle (heating to 50 °C followed by cooling to 20 °C), the heterogeneous coacervate exhibited an irreversible first-order phase transition to an anisotropic ion gel. This established a coacervate-ion gel phase diagram having a well-defined UCST.

Linear viscoelasticity of complex coacervates

“…In numerous examples, the elastic behavior dominates (i.e., the storage modulus (G') dominates) [8,61,137,138,171,175,[177][178][179][180], while in other coacervates the viscous or liquid-like behavior (i.e., the loss modulus (G") dominates) [95,103,171]. Meanwhile, there are other systems where a crossover is observed between the two regimes, with G" dominating at low frequencies, and G' dominating at higher frequencies [61,71,72,79,111,130,162,171].…”

“…Firstly, the application of temperature to many of the materials used in coacervation, and proteins in particular, can lead to a gelation phenomenon. In protein-based systems such as those involving gelatin, this type of gel formation is often the result of temperature or pH-induced protein denaturation [137,182]. Solid-to-liquid transitions such as gelation can be observed from a plot of tan(δ) versus frequency.…”

Coacervation in Biopolymers

“…It has been shown that a polyelectrolyte, DNA and a polyampholyte, gelatin can undergo associative interaction and form complex coacervates with interesting thermal properties [15]. Further, it has been realized that in a class of systems coacervation transition is governed by surface selective patch binding even though both the polyions carry similar net charge [11,18,19].…”

“…In particular, protein-polysaccharide and protein-protein coacervates have attracted much attention because of their inherent potential in generating new biomaterials. In addition, such studies provide basic understanding of specific and non-specific interactions operating between complementary polyelectrolytes [1-6] or polyelectrolyte-polyampholyte [7][8][9][10][11][12][13][14][15] pairs. Normally, polysaccharides are strong polyelectrolytes whereas proteins, in addition, can be polyampholytes.…”

“…In contrast, *Corresponding author: Kamla Rawat, Special Center for Nanosciences, Jawaharlal Nehru University, New Delhi 110067, India, Tel: +91 11 2670 4699; Fax: +91 11 2674 1837; E-mail: kamla.jnu@gmail.com β-lactoglobulin-pectin coacervates were found to be a heterogeneous phase comprising of pectin networks with protein domains forming the junction points [17]. It has been shown that a polyelectrolyte, DNA and a polyampholyte, gelatin can undergo associative interaction and form complex coacervates with interesting thermal properties [15]. Further, it has been realized that in a class of systems coacervation transition is governed by surface selective patch binding even though both the polyions carry similar net charge [11,18,19].…”