Abstract
Charged amino acids having ionizable side chains play crucial roles in maintaining the solubility and stability of a protein. These charged amino acids are mostly exposed on protein surface and participate in electrostatic interactions with neighboring charged amino acids as well as with solvent. Therefore, the change in the solvent pH affects the protein stability in most cases. Previously, we reported a GFP variant, GFP14R having 14 surface lysines replaced with arginines, that showed enhanced stability under alkaline pH. Here, we analyzed the factors that contribute to the stability of the GFP14R under alkaline pH quantitatively using molecular dynamics simulations. Protonation state of the charged amino acids of GFP14R and control GFP under neutral pH and alkaline pH were modeled, and molecular dynamics simulations were performed. This comparative analysis revealed that the GFP14R with more arginine frequency on the surface maintained the stability under both pH conditions without much change in their salt-bridge interactions as well as the hydrogen bond interactions with solvent. On the other hand, these interactions were significantly reduced for the control GFP under alkaline pH due to the deprotonated lysine side chains. These results suggest that the advantageous property of arginine over lysine can be considered one of the parameter for the protein stability engineering under alkaline pH conditions.
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Sokalingam, S., Madan, B., Raghunathan, G. et al. Deciphering the factors responsible for the stability of a GFP variant resistant to alkaline pH using molecular dynamics simulations. Biotechnol Bioproc E 18, 858–867 (2013). https://doi.org/10.1007/s12257-013-0309-1
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DOI: https://doi.org/10.1007/s12257-013-0309-1