An able-cryoprotectant and a moderate denaturant: distinctive character of ethylene glycol on protein stability

Abstract

Osmolytes are known to stabilize proteins against denaturing conditions. Ethylene glycol (EG), however, shows a distinctive effect on α-lactalbumin (α-LA) that it stabilizes the protein against cold-induced denaturation, whereas it destabilizes during heat denaturation. The replica exchange molecular dynamics (REMD) simulation of α-LA in the presence of EG shows that EG denatures the protein at higher temperatures whereas it retards the denaturation at sub-zero temperature. Representative structures of α-LA were selected from REMD trajectories at three different temperature conditions (240, 300 and 340 K) with and without EG, and classical molecular dynamics (MD) simulations were performed. The results suggest that the presence of water around α-LA is more at lower temperatures; however, water around the hydrophobic residues is reduced with the addition of EG at sub-zero temperature. The partition coefficient of EG showed that the binding of EG with hydrophobic residues was higher at lower temperatures. Preferential interaction parameters at different temperatures were calculated based on the mean distribution (Γ₂₃) and Kirkwood–Buff integral (G₂₃) methods. Γ₂₃ shows a larger positive value at 240 K compared to higher temperatures. G₂₃ shows positive values at lower temperatures, whereas it becomes negative at above 280 K. These results indicate that the preferential binding of EG with α-LA is more at sub-zero temperature compared to higher temperature conditions. Thus, the study suggests that the preferential binding of EG reduces the hydrophobic hydration of α-LA at lower temperatures, and stabilizes the protein against cold denaturation. However, the preferential binding of EG at higher temperature drives the folding equilibrium towards the denatured state.

Communicated by Ramaswamy H. Sarma

Keywords:

• Cold denaturation of protein
• thermal denaturation of protein
• replica exchange molecular dynamics simulation
• α-lactalbumin
• ethylene glycol

Acknowledgements

The authors thank CSIR, India (01 (2845)/16/EMR-11) and SERB, India (EMR/2016/003411) for the financial support. The authors also thank CMSD and BIF at University of Hyderabad for computational facilities and DST-FIST (SR/FST/LSI-530/2012) and UGC-SAP (SAP-198-2014-108) grants for instrumentation facilities.

Disclosure statement

There are no conflicts of interest to declare.

Additional information

Funding

DST-FIST (SR/FST/LSI-530/2012) and UGC-SAP (SAP-198-2014-108).