QUESTION 1. a) You are studying the protein Hrd1, which has been implicated in r

Question

QUESTION 1. a) You are studying the protein Hrd1, which has been implicated in rheumatoid arthritis. Hrd1 is a multi- pass transmembrane protein that detects misfolded proteins as part of the “unfolded protein response" and tags them with Ubiquitin. One of the misfolded proteins that it can ubiquitinate is called MHC1. The functional Hrd1 protein has 6 transmembrane domains, but it lacks the traditional N-terminal ER signal sequence. If you performed immunocytochemistry with an antibody to Hrd1 protein, where in the cell would most of the signal be located? (2 pts) b) Draw the orientation of this protein in an organelle within the endomembrane system, being sure to denote the N-terminus ("N"), C-terminus ("C"), membrane, endomembrane lumen, and cytosol. (2 pts) c) (4 pts) There are 2 asparagines in the Hrd1 protein, one facing the endomembrane lumen (at amino acid 300) and one facing the cytosol (at amino acid 550). You have two different mutant cell culture lines in which each has a different single asparagine-to-alanine substitution in your Hrd1 protein. Based on what we learned in class about asparagine residues, which mutant version is more likely to have defective Hrd1 activity (circle), and why? (4 pts) The one with mutation at amino acid 300 The one with mutation at amino acid 550

Explanation / Answer

1A) As protein contains transmembrane domains and known to ubiquitinate other proteins most likely immunostaining will reveal its location on Golgi bodies of the cell.

1C) Aspargine amino acid residue has a side chain available or hydrogen bonding, therefore it is present mostly at the begining or end of alpha helix slices or turn of beta sheets. Mutation occurring due aspargine getting substituted with alanine at cytosolic end is more likely to hamper protein activity by misfolding. Hence mutation at 550aa position is more likely to hamper its function. Second mutant likely is more likely to have hampered Hrd1 protein activity.

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