Salt-Bridges in the Microenvironment of Stable Protein Structures

Abstract

Salt-bridges (sb) play an important role in the folding and stability of proteins. This is deduced from the evaluation of net energy in the microenvironments (ME, residues that are 4Å away from positive and negative partners of salt-bridge and interact with them). ME’s act as a determinant of net-energy due to the intrinsic features by the sequence. The stability of extremophilic proteins is due to the presence of favorable residues at the ME without any unfavorable residues. We studied a dataset of four structures from the pdb and a homology model (PDB ID: 1HM5) to gain insights on this issue. Data shows that the presence of isolated charges and polar residues in the core of extremophilic proteins helps in the formation of stable salt-bridges with reduced desolvation. Thus, site-specific mutations with favorable residues at the ME will help develop thermo stable proteins with strong salt bridges.

Intrinsic basis of thermostability of prolyl oligopeptidase from Pyrococcus furiosus

“…It needs to be mentioned here that, in order to have a prominent global effect, ME-population needs to have the most adverse candidates. Compared to other studies 53 HuP's ME-population does not have such a candidate in it, and thus, the global effect, although favorable, is less prominent. Although the replacement of our ME-residue stabilizes the unstable salt-bridge, the limitations of the in-silico method need to be kept in mind here.…”

“…Thus, for a given salt-bridge, preparation of five different mutated PDB structures and nine different APBS runs were required to obtain the component energy terms 39 , 63 – 65 . A higher version of the earlier programs 63 – 65 was used for automated extraction of the component and hence the net energy terms 53 .…”

“…The side chains of the partners of a salt-bridge could be surrounded by isolated charged, polar and hydrophobic residues of a protein. These residues influence the background and hence, the net energy terms of a salt-bridge 36 – 39 , 53 . When all salt-bridges of a protein are considered, we obtain a collection of these residues.…”

“…A residue is taken as ME of a salt-bridge, when the energy of interaction between the partners of a salt-bridge and the residue is lower than (stabilizing) or higher than (destabilizing) a preset energy cut-off. We set the cut-off at 0.75 kJ/mol 53 . We then raised the following questions.…”

“…Unfavorable (positive interaction-energy) and favorable (negative interaction-energy) ME-residues at different locations (core or surface) and in a segment of secondary structure (helix or strand or coil) of protein could be identified. Mutation of the targeted ME-residue was performed in Swiss-Pdbviewer, v4.1.0 53 , 66 by using the lowest steric clashed (with the main and side chains of neighboring residues) rotamer of the residue. The minimized structure of a protein was used for the targeted mutation for a highly unfavorable or favorable residue.…”