A general summation of the question is as follows:
I am "performing" an analysis of membrane proteins based on the separation between aqueous and detergent phase (using Triton X-114). Membranes are made soluble with the detergent at 4 degrees C, then the temp is changed to 30 degrees C to separate proteins based on solubility into either detergent or aqueous phase. I then "conduct" two gel electrophoresis samples; one using coomassie blue, and the other using a galactose specific stain. I have a photo of the two gels showing the protein bands at various points (not sure if I need to post the photo or not).
The questions are:
1) What bands correspond to transmembrane proteins, and what bands to peripheral proteins (using coomassie blue stain gel).
2) What proteins are glycosylated, and what is the relationship to those in the first gel? (analysis of the galactose-specific stain)
1) Coomassie Blue stain gel contains three wells to show total membrane, aqueous phase, and detergent phase.
Aqueous phase should show me hydrophilic protiens (peripheral proteins?) and detergent phase should show me integral membrane proteins (transmembrane proteins?).
2) The Galactose staining pattern (gel) again shows the same three wells as the first gel, but different bands (and band widths)
The galactose stain should show proteins that have been glycosylated... but what am I supposed to conclude about these glycosylated proteins to the integrated/peripheral proteins? Aren't glycosylated proteins only exposed to the inner/outer atmosphere of the cell? (AKA: not within the lipid bilayer) So why am I seeing bands that are present on the integrated proteins?
Unless it's saying that it's a transmembrane protein, so the edge that is 'exposed' is capable of being glycosylated and therefore would show up on the stain...
Any help? I can post the photo if needed.
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