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Unformatted text preview: . v-Src is attached to the membrane via a myristoyl
group linked to its N-terminal glycine. Proteins can also be palmitoylated allowing
them to attach to the membrane. They can be purified by treating cells with
Oligosaccharide linkage to phosphatidylinositol: e.g. Thy-1 is attached via a GPI
(glycosylphosphatidylinositiol) linkage. Such proteins are usually only found on the
extracellular side of the membrane. It can be purified by treating with a PI-specific
Peripheral protein: e.g. Phospholipase A2. It can be extracted by breaking up the
electrostatic interactions using high salt concentrations or a high pH. Some
peripheral membrane proteins need calcium ions to form electrostatic interactions 7.06 Spring 2004 PS 1 key 10 of 11 with the phospholipid head groups and so these proteins can be released by adding a
(b) Why is it that polypeptides that enter the bilayer tend to pass completely through it
rather than changing direction midway?
Changing direction midway would involve making a bend in the polypeptide
backbone, which is not energetically favored as it decreases the number of hydrogen
bonds formed. Therefore, from a thermodynamic viewpoint it is much better if a
protein passes through the entire bilayer.
(c) You have cloned a multipass transmembrane receptor and are trying to determine
whether the N-terminus is on the cytoplasmic or the extracellular side of the bilayer.
How would the following information help you determine this:
position of glycosylated residues
a series of monoclonal antibodies whose epitopes have been
(iii) a recombinant form of the protein that has been tagged at the
(i) Glycosylated residues are always found on the extracellular portion of a
transmembrane protein – in a future lecture on protein secretion and modification
we will discover why this is.
(ii) If you have a monoclonal antibody you could incubate cells that express the
receptor with this antibody, wash away any unbound antibody, cross-link bound
antibody and then add a secondary antibody coupled to a fluorescent molecule.
Antibodies can’t diffuse thro...
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- Spring '11