Which terminal is extracellular and intracellular who is?
Fluorescence microscopy can be used to study protein topology. In one experiment, constructs were designed to express two membrane proteins: the glycosylphosphatidylinositol (GPI) anchored protein prion, PrP, with yellow fluorescent protein (YFP) attached to its N-terminal (YFP-PrP) and HLAA2 with cyan fluorescent protein (CFP) attached at its C-terminal (HLA-A2-CFP). The cells were imaged expressing these fusion proteins in the presence and absence of trypsin. Given these results in the image below. http://yfrog.com/55ps3qp What can you conclude about the topology of membrane proteins? (That say which terminal is extracellular and intracellular who is?)
To answer this question, it will help you know what the trypsin protein. Trypsin split clumps of cells by degrading proteins that bind them. It does so by cutting the main peptide chains on the carboxyl side of amino acids lysine or arginine. A cluster of cells are linked to each other by proteins that are integral to the extracellular face (tight junctions) or by proteins facing integral intracellular (junctions). These proteins are cleaved when cells are split because extracellular trypsin is too large to enter the cells so intracellular proteins are not affected (. now with this knowledge, you can see in the image you provided, the green / yellow is extremely decreased between - Trypsin) and (+ trypsin), while red is the same. The issues mentioned that YFP is attached to the N-terminus and CFP is attached to the C-terminus. This means that the N-terminal must be outside and is degraded by trypsin. NC-extracellular intracellular
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