6. You are studying a cargo protein which you believe is involved in nuclear tra
ID: 262884 • Letter: 6
Question
6. You are studying a cargo protein which you believe is involved in nuclear transport. You have created various truncation mutations in the cargo protein in which certain portions of are removed. You determine whether the cargo protein localizes to the nucleus and/or cytoplasm under normal conditions and also in the presence of leptomycin B, an inhibitor of the nuclear export machinery Your data are shown in the following figure.(A plus means the cargo protein is located in that part of the cell, a minus that it isn't.) 151.750 301-75o 451-750 01-430 51-450 1-150 151-300 A. (4pts) Why does the removal of parts of this cargo protein (and others) cause the cargo protein to change its location in the cell? B. (4pts) How can you use this information to determine the location of a nuclear localization sequence (NLS) in this cargo protein? C. (2pts) What range of amino acids do you think contains the NLS? D. (2pts) What range of amino acids do you think contains the NES?Explanation / Answer
A) Removal of parts of the cargo protein may lead to the removal of the binding site (the nuclear localization sequence or the nuclear export signals) required by the importin/exportin to bind to the cargo so as to transport the protein from the nucleus to the cytoplasm (or vice-versa). If the exportin cannot bind to the cargo, it stays in the nucleus, whereas if it can bind to the cargo, it is transported to the cytoplasm. Similarly, if the importin can bind to the protein, is can be transported to the nucleus from the cytoplasm, otherwise it cannot.
B) By comparing the cases with different cargo protein lengths, we can see that in cases where there is protein in the cytoplasm but not in the nucleus (in both conditions), the import of protein from the cytoplasm to the nucleus is not occurring which means that the NLS is missing. This can be found by comparing the cases with import and no import.
C) 301-450 and 1-150 lengths contain the NLS.
D) 151-300 and 451-750 lengths contain the NES.
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