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#protein orientation

2 public questions tagged with this topic.

Plasma Membrane Proteins

Which technique can be used to determine if a membrane protein is inside-out or right-side-out?

Determining orientation of membrane proteins after reconstitution into artificial vesicles essential for establishing vectorial transport competence. Proteoliposomes generated by removing detergent via dialysis may incorporate proteins randomly right-side-out exposing original extracellular domains outward or inside-out exposing cytoplasmic domains outward leading to mixed activity. Sidedness assay exploits membrane impermeability of antibodies one hundred fifty kilodaltons IgG unable to cross sealed bilayer. Intact vesicles incubated with monoclonal antibody against known extracellular epitop

Ref: Rohde et al., Determination of Membrane Protein Orientation by Antibody Labeling, J Mol Biol Methods.

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Protein Sorting and Trafficking

The topology of membrane proteins is determined by:

Topological orientation of membrane proteins, defined by which loops face cytosolic versus exoplasmic or lumenal space, is encoded within polypeptide itself as a set of topogenic signals rather than imposed solely by external receptors. As nascent chain enters Sec61 translocon, hydrophobic segments can function either as start-transfer signal-anchor that opens lateral gate and initiates translocation of downstream region, or as stop-transfer sequence that halts translocation and partitions laterally into lipid bilayer through same gate formed by helices 2b and 7. Positive-inside rule, first do

Ref: von Heijne G, Annu Rev Biophys 36: 2007, Membrane Protein Topology Determinants.

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