Kinetic stability of membrane proteins

Abstract

Although membrane proteins constitute an important class of biomolecules involved in key cellular processes, study of the thermodynamic and kinetic stability of their structures is far behind that of soluble proteins. It is known that many membrane proteins become unstable when removed by detergent extraction from the lipid environment. In addition, most of them undergo irreversible denaturation, even under mild experimental conditions. This process was found to be associated with partial unfolding of the polypeptide chain exposing hydrophobic regions to water, and it was proposed that the formation of kinetically trapped conformations could be involved. In this review, we will describe some of the efforts toward understanding the irreversible inactivation of membrane proteins. Furthermore, its modulation by phospholipids, ligands, and temperature will be herein discussed.

An internally quenched peptide as a new model substrate for rhomboid intramembrane proteases

“…However, the inherent hydrophobic nature of intramembrane proteases comes with particular challenges when in vitro work is undertaken. While most membrane proteins retain their stability in detergent solutions, there are many examples where they become unstable when removed from the lipid environment by detergent extraction (Gonzalez Flecha, 2017). In addition, some membrane proteins have preferences for special detergents or lipids.…”

Conjugative Coupling Proteins and the Role of Their Domains in Conjugation, Secondary Structure and in vivo Subcellular Location

“…This result suggests that although the secondary structures of the mutants differ from that of the native protein, their overall thermal stability is similar. Additionally, when comparing the results between both studied systems, as expected due to the high instability of purified membrane proteins ( González Flecha, 2017 ) the soluble proteins showed higher mid-point denaturation temperatures than the full-length proteins. Specifically, MobBΔTMD and TrwBΔN70 showed similar Tm values (Tm 62 and 63°C, respectively); on the contrary, MobB CloDF13 was more stable than TrwB R388 against thermal denaturation (Tm 56 and 48°C, respectively).…”

Unfolding of Helical Poly(L-Glutamic Acid) in N,N-Dimethylformamide Probed by Pyrene Excimer Fluorescence (PEF)

“…The stability of a protein is usually defined by its ability to resist unfolding upon being subject to denaturing forces resulting from the addition of a denaturing agent, a sudden change in solution pH, or a step increase in temperature [ 28 , 29 , 34 ]. A quantitative measure of the stability of a protein is provided by the change in Gibbs energy (Δ unfold G (H 2 O)) between the folded ( N ) and fully unfolded ( D ) conformation of a protein in water.…”

“…The N blob value was also employed to determine the molar fraction of native ( f N ) and denatured ( f D ) P L GA molecules in solution. In turn, the f N and f D fractions could be applied to determine the equilibrium constant ( K unfold ) for the Native ⇌ Denatured equilibrium at each GdHCl concentration [ 28 , 29 ] and yield the change in Gibbs energy (Δ unfold G (DMF)) for the unfolding of the α–helical P L GA in DMF in the same manner, that experiments using CD or LS would do. In summary, these PEF experiments further support the notion that N blob , determined for macromolecules randomly labeled with pyrene, reports on the local density of macromolecules and can be used to infer their conformation in solution.…”