Electrostatic contribution to the thermodynamic and kinetic stability of the homotrimeric coiled coil Lpp‐56: A computational study

Abstract

Electrostatic contribution to the thermodynamic and kinetic stability of the homotrimeric coiled coil Lpp-56: A computational study 2006;45:8931-8939). The unfolding rate constant measured using GdmCl as the denaturing agent, though extremely low, was substantially higher than that obtained on the basis of thermal unfolding. If this large difference arises from the effect of screening of electrostatic interactions induced by GdmCl, electrostatic interactions would appear to be an important factor determining the unusual properties of Lpp-56. We present here a computational analysis of the electrostatic properties of Lpp-56 combining molecular dynamics simulations and continuum pK calculations. The pH-dependence of the unfolding free energy is predicted in good agreement with the experimental data: the change in DeltaG between pH 3 and pH 7 is approximately 60 kJ mol(-1). The results suggest that the difference in the stability of the protein observed using different experimental methods is mainly because of the effect of the reduction of electrostatic interactions when the salt (GdmCl) concentration increases. We also find that the occupancy of the interhelical salt bridges is unusually high. We hypothesize that electrostatic interactions, and the interhelical salt bridges in particular, are an important factor determining the low unfolding rate of Lpp-56. is an attractive object of biophysical investigation in several aspects. It is a homotrimeric, parallel coiled coil, a class of coiled coils whose stability and folding have been studied only occasionally. Lpp-56 possesses unique structural properties and exhibits extremely low rates of folding and unfolding. It is natural to ask how the specificity of the structure determines the extraordinary physical chemical properties of interactions would appear to be an important factor determining the unusual properties of Lpp-56. We present here a computational analysis of the electrostatic properties of Lpp-56 combining molecular dynamics simulations and continuum pK calculations. The results suggest that the difference in the stability of the protein observed using different experimental methods is mainly due to effect of the reduction of electrostatic interactions when the salt (GdmCl) concentration increases. We also find that the occupancy of the interhelical salt bridges is unusually high. We hypothesize that electrostatic interactions, and the interhelical salt bridges in particular, are an important factor determining the low unfolding rate of Lpp-56.

Shedding light on the extra thermal stability of thermophilic proteins

“…In fact, such ion pairs and charge–charge networks can provide an efficient way to couple together different structural regions of the protein, producing low‐frequency (ie, long wavelength) vibrational modes. Note, in this respect, that the performed computational studies have not detected a clear energetic stabilization afforded by the ion pairs and charge–charge networks . Such a coupling mechanism, involving exposed and charged side‐chains, can provide an explanation for the abundance of low‐frequency vibrational modes and their significant population also at room temperature.…”

The L49F mutation in alpha erythroid spectrin induces local disorder in the tetramer association region: Fluorescence and molecular dynamics studies of free and bound alpha spectrin

“…A comparison of currently available force fields show different structural features for different force fields, and no single force field is available for all structures 25. Although some force fields favor α‐helix and others favor β‐hairpin, OPLS‐AA/L, the force field used in this study, has intermediate tendency,25 and has been applied to coiled coil structures 26. Using an identical simulation protocol, we were able to compare differences between WT and L49F.…”

Rigidity versus flexibility: the dilemma of understanding protein thermal stability

“…It contains six rings of salt bridges tightening the coiled coil, in this way making an important contribution to the electrostatic term of the free energy. This feature of the protein was revealed after MD simulation , whilst use of the X‐ray structure (providing fixed atomic coordinates) led to a dramatic underestimation of the electrostatic contribution to the folding free energy. An extraordinary property of Lpp‐56, explained by the long lifetime of the salt bridge networks, is its extremely low unfolding rate (10 −9 – 10 −11 s −1 ) .…”