Influence of Pathogenic Mutations on the Energetics of Translocon-Mediated Bilayer Integration of Transmembrane Helices

Schlebach, Jonathan P.; Sanders, Charles R.

doi:10.1007/s00232-014-9726-0

Cited by 23 publications

(31 citation statements)

References 41 publications

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“…Previously reported motor nerve conduction velocities of patients harboring these mutations confirm that the degree of dysmyelination caused by these mutations is highly variable ( Supplementary Table 1 ), which suggests that this set of mutations is likely to manifest a range of effects on the conformational equilibrium of PMP22. Additionally, we recently found that none of these mutations are predicted to cause topogenic defects, 23 which suggests that conformational defects are manifested after PMP22 is cotranslationally inserted into the membrane.…”

Section: Resultsmentioning

confidence: 99%

Conformational Stability and Pathogenic Misfolding of the Integral Membrane Protein PMP22

Schlebach¹,

Narayan²,

Alford

et al. 2015

J. Am. Chem. Soc.

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138

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Despite broad biochemical relevance, our understanding of the physiochemical reactions that limit the assembly and cellular trafficking of integral membrane proteins remains superficial. In this work, we report the first experimental assessment of the relationship between the conformational stability of a eukaryotic membrane protein and the degree to which it is retained by cellular quality control in the secretory pathway. We quantitatively assessed both the conformational equilibrium and cellular trafficking of 12 variants of the α-helical membrane protein peripheral myelin protein 22 (PMP22), the intracellular misfolding of which is known to cause peripheral neuropathies associated with Charcot–Marie–Tooth disease (CMT). We show that the extent to which these mutations influence the energetics of Zn(II)-mediated PMP22 folding is proportional to the observed reduction in cellular trafficking efficiency. Strikingly, quantitative analyses also reveal that the reduction of motor nerve conduction velocities in affected patients is proportional to the extent of the mutagenic destabilization. This finding provides compelling evidence that the effects of these mutations on the energetics of PMP22 folding lie at the heart of the molecular basis of CMT. These findings highlight conformational stability as a key factor governing membrane protein biogenesis and suggest novel therapeutic strategies for CMT.

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Section: Resultsmentioning

confidence: 99%

Conformational Stability and Pathogenic Misfolding of the Integral Membrane Protein PMP22

Schlebach¹,

Narayan²,

Alford

et al. 2015

J. Am. Chem. Soc.

Self Cite

138

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“…Even single mutations to an IMP amino-acid sequence can disrupt integration and induce disease phenotypes [16] or decrease protein expression [17–19]; similarly, mutations to the translocon channel can inhibit IMP folding [8, 20–23]. The important role for IMPs in cellular functions, such as signal transduction, the transport of nutrients, and cell adhesion, motivates the understanding of the effect of NC and translocon properties on the efficiency of co-translational integration.…”

Section: Introductionmentioning

confidence: 99%

Structurally detailed coarse-grained model for Sec-facilitated co-translational protein translocation and membrane integration

et al. 2017

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We present a coarse-grained simulation model that is capable of simulating the minute-timescale dynamics of protein translocation and membrane integration via the Sec translocon, while retaining sufficient chemical and structural detail to capture many of the sequence-specific interactions that drive these processes. The model includes accurate geometric representations of the ribosome and Sec translocon, obtained directly from experimental structures, and interactions parameterized from nearly 200 μs of residue-based coarse-grained molecular dynamics simulations. A protocol for mapping amino-acid sequences to coarse-grained beads enables the direct simulation of trajectories for the co-translational insertion of arbitrary polypeptide sequences into the Sec translocon. The model reproduces experimentally observed features of membrane protein integration, including the efficiency with which polypeptide domains integrate into the membrane, the variation in integration efficiency upon single amino-acid mutations, and the orientation of transmembrane domains. The central advantage of the model is that it connects sequence-level protein features to biological observables and timescales, enabling direct simulation for the mechanistic analysis of co-translational integration and for the engineering of membrane proteins with enhanced membrane integration efficiency.

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“…Establishing the principles governing the folding of a membrane protein is central to understanding the molecular basis for membrane proteins that display multiple topologies and a wide spectrum of membrane protein conformational and topological disorders (1)(2)(3)(4)(5)(6). At least 10% of 470 known pathogenic mutations were predicted to result in a change in the topological organization of the mutant membrane protein (7). A fundamental objective in membrane biology is to understand and predict how protein sequence determines the number and orientation of transmembrane domains (TMDs) 3 (8).…”

mentioning

confidence: 99%

Dynamic Lipid-dependent Modulation of Protein Topology by Post-translational Phosphorylation

Vitrac¹,

MacLean²,

Karlstaedt

et al. 2017

Journal of Biological Chemistry

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Membrane protein topology and folding are governed by structural principles and topogenic signals that are recognized and decoded by the protein insertion and translocation machineries at the time of initial membrane insertion and folding. We previously demonstrated that the lipid environment is also a determinant of initial protein topology, which is dynamically responsive to post-assembly changes in membrane lipid composition. However, the effect on protein topology of post-assembly phosphorylation of amino acids localized within initially cytoplasmically oriented extramembrane domains has never been investigated. Here, we show in a controlled in vitro system that phosphorylation of a membrane protein can trigger a change in topological arrangement. The rate of change occurred on a scale of seconds, comparable with the rates observed upon changes in the protein lipid environment. The rate and extent of topological rearrangement were dependent on the charges of extramembrane domains and the lipid bilayer surface. Using model membranes mimicking the lipid compositions of eukaryotic organelles, we determined that anionic lipids, cholesterol, sphingomyelin, and membrane fluidity play critical roles in these processes. Our results demonstrate how post-translational modifications may influence membrane protein topology in a lipid-dependent manner, both along the organelle trafficking pathway and at their final destination. The results provide further evidence that membrane protein topology is dynamic, integrating for the first time the effect of changes in lipid composition and regulators of cellular processes. The discovery of a new topology regulatory mechanism opens additional avenues for understanding unexplored structure-function relationships and the development of optimized topology prediction tools.

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Influence of Pathogenic Mutations on the Energetics of Translocon-Mediated Bilayer Integration of Transmembrane Helices

Cited by 23 publications

References 41 publications

Conformational Stability and Pathogenic Misfolding of the Integral Membrane Protein PMP22

Conformational Stability and Pathogenic Misfolding of the Integral Membrane Protein PMP22

Structurally detailed coarse-grained model for Sec-facilitated co-translational protein translocation and membrane integration

Dynamic Lipid-dependent Modulation of Protein Topology by Post-translational Phosphorylation

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