Effect of osmolytes on the binding of EGR1 transcription factor to DNA

Abstract

Osmolytes play a key role in maintaining protein stability and mediating macromolecular interactions within the intracellular environment of the cell. Herein, we show that osmolytes such as glycerol, sucrose and PEG400 mitigate the binding of EGR1 transcription factor to DNA in a differential manner. Thus, while physiological concentrations of glycerol only moderately reduce the binding affinity, addition of sucrose and PEG400 is concomitant with a loss in the binding affinity by an order of magnitude. This salient observation suggests that EGR1 is most likely subject to conformational equilibrium and that the osmolytes exert their effect via favorable interactions with the unliganded conformation. Consistent with this notion, our analysis reveals that while EGR1 displays rather high structural stability in complex with DNA, the unliganded conformation becomes significantly destabilized in solution. In particular, while liganded EGR1 adopts a well-defined arc-like architecture, the unliganded protein samples a comparatively large conformational space between two distinct states that periodically interconvert between an elongated rod-like shape and an arc-like conformation on a sub-microsecond time scale. Consequently, the ability of osmolytes to favorably interact with the unliganded conformation so as to stabilize it could account for the negative effect of osmotic stress on EGR1-DNA interaction observed here. Taken together, our study sheds new light on the role of osmolytes in modulating a key protein-DNA interaction.

Osmotic modulation of chromatin impacts on efficiency and kinetics of cell fate modulation

“…The most interesting effects of the hyposmotic condition were seen in the gene regulatory networks of EGR1, EGR3, Jun and ATF4. The presence of EGR1 was an interesting hit because it is reported in the literature as having activity impairment with increasing amounts of osmolytes 41 . One of the first events after the hyposmotic stimulus is the exit of osmolytes from the cell to balance the extracellular hyposmotic environment.…”

Exploring conformational preferences of proteins: ionic liquid effects on the energy landscape of avidin

“…All ILs were characterised by 1 H and 13 C NMR spectroscopy in either DMSO-d 6 or D 2 O at 25 C using Bruker Advance III, 400 MHz spectrometer (Bruker Corporation, Billerica, MA, USA). Electrospray ionisation mass spectrometry was performed using Waters LCT Premier Mass Spectrometer (Waters Corporation, Milford, MA, USA) in positive and negative ion mode.…”

“…2,[10][11][12] The goal to determine which selective pressures can induce a stable protein conformation is further complicated by the large conformational space a protein can sample, preferential interactions available, and environmental conditions. 13 This begs the question: Can we tune environmental factors to select for more stable protein conformations?…”

Biological Wonders of Osmolytes: The Need to Know More

“…The destabilization of DNA by osmolytes has also been observed [79] and role in modulating protein-DNA interactions has been discussed [80]. For example, mitigation of the binding of ERG1 to DNA in a differential manner has been reported.…”