2. Trypsin is a hydrophilic protease and therefore is unable to cross the plasma

Question

2. Trypsin is a hydrophilic protease and therefore is unable to cross the plasma membrane and enter a cell. However, it is capable of digesting the hydrophilic portions of membrane proteins. As a result of these properties, researchers investigating membrane proteins have treated cells with trypsin then separated the proteins by SDS-PAGE to gain insight into the orientation of particular proteins within the membrane. Describe an experiment using this information and the results you would expect with the following types of membrane-associated proteins. (Assume the proteins are single polypeptides.) a. an integral membrane protein with the N-terminus exposed on the surface b. an integral membrane protein with the C-terminus exposed on the surface c. an integral membrane protein with three hydrophilic loops (only) exposed on the surface d. a membrane-associated protein facing the cytoplasm e. an imbedded membrane protein

Explanation / Answer

SDS-PAGE is a systemetic technique for proteins separation on the basis of their molecular weight. Smaller fragments migrate faster due to less resistance as compared to larger fragments. In SDS-PAGE, sodium dodecyl sulfate (SDS) as well as polyacrylamide gel are used to reduces the effects of the structure and charge, and proteins got separated only based on the chain length of polypeptide. we would expect the result after digesting with trypsin as follows:

c. consist of 2-4 fragments
d. 1 fragment
e. 1 fragment

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