TY - JOUR
T1 - Cys-labelling kinetics of membrane protein GlpG
T2 - a role for specific SDS binding and micelle changes?
AU - Otzen, Daniel E
AU - Pedersen, Jannik Nedergaard
AU - Somavarapu, Arun Kumar
AU - Clement, Anders
AU - Ji, Ming
AU - Petersen, Emil Hartvig
AU - Pedersen, Jan Skov
AU - Urban, Sinisa
AU - Schafer, Nicholas P
N1 - Copyright © 2021 Biophysical Society. Published by Elsevier Inc. All rights reserved.
PY - 2021/9
Y1 - 2021/9
N2 - Empirically, α-helical membrane protein folding stability in surfactant micelles can be tuned by varying the mole fraction MF
SDS of anionic (sodium dodecyl sulfate (SDS)) relative to nonionic (e.g., dodecyl maltoside (DDM)) surfactant, but we lack a satisfying physical explanation of this phenomenon. Cysteine labeling (CL) has thus far only been used to study the topology of membrane proteins, not their stability or folding behavior. Here, we use CL to investigate membrane protein folding in mixed DDM-SDS micelles. Labeling kinetics of the intramembrane protease GlpG are consistent with simple two-state unfolding-and-exchange rates for seven single-Cys GlpG variants over most of the explored MF
SDS range, along with exchange from the native state at low MF
SDS (which inconveniently precludes measurement of unfolding kinetics under native conditions). However, for two mutants, labeling rates decline with MF
SDS at 0–0.2 MF
SDS (i.e., native conditions). Thus, an increase in MF
SDS seems to be a protective factor for these two positions, but not for the five others. We propose different scenarios to explain this and find the most plausible ones to involve preferential binding of SDS monomers to the site of CL (based on computational simulations) along with changes in size and shape of the mixed micelle with changing MF
SDS (based on SAXS studies). These nonlinear impacts on protein stability highlights a multifaceted role for SDS in membrane protein denaturation, involving both direct interactions of monomeric SDS and changes in micelle size and shape along with the general effects on protein stability of changes in micelle composition.
AB - Empirically, α-helical membrane protein folding stability in surfactant micelles can be tuned by varying the mole fraction MF
SDS of anionic (sodium dodecyl sulfate (SDS)) relative to nonionic (e.g., dodecyl maltoside (DDM)) surfactant, but we lack a satisfying physical explanation of this phenomenon. Cysteine labeling (CL) has thus far only been used to study the topology of membrane proteins, not their stability or folding behavior. Here, we use CL to investigate membrane protein folding in mixed DDM-SDS micelles. Labeling kinetics of the intramembrane protease GlpG are consistent with simple two-state unfolding-and-exchange rates for seven single-Cys GlpG variants over most of the explored MF
SDS range, along with exchange from the native state at low MF
SDS (which inconveniently precludes measurement of unfolding kinetics under native conditions). However, for two mutants, labeling rates decline with MF
SDS at 0–0.2 MF
SDS (i.e., native conditions). Thus, an increase in MF
SDS seems to be a protective factor for these two positions, but not for the five others. We propose different scenarios to explain this and find the most plausible ones to involve preferential binding of SDS monomers to the site of CL (based on computational simulations) along with changes in size and shape of the mixed micelle with changing MF
SDS (based on SAXS studies). These nonlinear impacts on protein stability highlights a multifaceted role for SDS in membrane protein denaturation, involving both direct interactions of monomeric SDS and changes in micelle size and shape along with the general effects on protein stability of changes in micelle composition.
U2 - 10.1016/j.bpj.2021.08.001
DO - 10.1016/j.bpj.2021.08.001
M3 - Journal article
C2 - 34370995
SN - 0006-3495
VL - 120
SP - 4115
EP - 4128
JO - Biophysical Journal
JF - Biophysical Journal
IS - 18
ER -