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Proc Natl Acad Sci U S A. 2018 Aug 28;115(35):E8172-E8180. doi: 10.1073/pnas.1807981115. Epub 2018 Aug 13.

Proteomic analysis of monolayer-integrated proteins on lipid droplets identifies amphipathic interfacial α-helical membrane anchors.

Author information

1
Department of Biochemistry, Stanford University, Stanford, CA 94305.
2
Department of Structural Biology, Stanford University, Stanford, CA 94305.
3
Department of Chemical and Systems Biology, Stanford University, Stanford, CA 94305.
4
Department of Statistics, Stanford University, Stanford, CA 94305.
5
Department of Biology, Stanford University, Stanford, CA 94305 kopito@stanford.edu.

Abstract

Despite not spanning phospholipid bilayers, monotopic integral proteins (MIPs) play critical roles in organizing biochemical reactions on membrane surfaces. Defining the structural basis by which these proteins are anchored to membranes has been hampered by the paucity of unambiguously identified MIPs and a lack of computational tools that accurately distinguish monolayer-integrating motifs from bilayer-spanning transmembrane domains (TMDs). We used quantitative proteomics and statistical modeling to identify 87 high-confidence candidate MIPs in lipid droplets, including 21 proteins with predicted TMDs that cannot be accommodated in these monolayer-enveloped organelles. Systematic cysteine-scanning mutagenesis showed the predicted TMD of one candidate MIP, DHRS3, to be a partially buried amphipathic α-helix in both lipid droplet monolayers and the cytoplasmic leaflet of endoplasmic reticulum membrane bilayers. Coarse-grained molecular dynamics simulations support these observations, suggesting that this helix is most stable at the solvent-membrane interface. The simulations also predicted similar interfacial amphipathic helices when applied to seven additional MIPs from our dataset. Our findings suggest that interfacial helices may be a common motif by which MIPs are integrated into membranes, and provide high-throughput methods to identify and study MIPs.

KEYWORDS:

lipid droplets; lipid monolayer; membrane insertion; proteomics

PMID:
30104359
PMCID:
PMC6126764
[Available on 2019-02-28]
DOI:
10.1073/pnas.1807981115
[Indexed for MEDLINE]

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