Studying
Cell Biology Preface | Introduction to Problem Solving | Problem Sets | Acknowledgments |
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The
current model of the plasma membrane emphasizes its asymmetric properties,
in terms of both structure and function. How were these asymmetric
features established? Consider this question carefully, in light of
what you have learned so far this term and ALSO in light of the following
data, and answer a., b., c., d., and e. below. i. Exposed tyrosine residues of membrane proteins can be labeled with
125 I and membrane carbohydrate that have first been oxidized
can be labeled by reduction with tritiated borohydride (3H-BH4).
Both these reactions are catalyzed by enzymes that cannot cross the plasma
membrane. ii. When applied to intact mammalian erythrocytes, and the proteins extracted
and analyzed by SDS-PAGE, the same bands are labeled by each technique.
These represent 5 of the 20 or so major bands evident when the gel is
stained for protein. Two of the 5 bands are glycophorin and the
anion transporter (Band 3). iii.All 5 bands are also stained by the Periodic Acid-Schiff reaction
(PAS+); none of the PAS-negative bands is stained by either reaction. iv. Detailed sequence analysis of glycophorin indicates the sites of
both 3H and 125I labeling are located near the amino
terminal end of the protein. Similar analysis of Band III presents
a more complex picture: 125I is found at several, periodically
separated sites some distance from either the amino or carboxy terminal
ends; these sites are between 70 and 120 amino acids apart. Carbohydrate
labeled with 3H in Band 3 is found exclusively at one location
about 50 amino acids from the carboxy terminus. v. When these labeling reactions are carried out on the cytoplasmic side
of resealed ghosts, virtually all SDS-PAGE bands are labeled with 125I;
none is labeled with 3H-BH4 or stained by PAS. i.If the Na+/K+-pump were labeled by tritiated boron hydride, explain briefly why the label might not have been evident in a discrete SDS-PAGE band and why the labeling might not have inhibited the pump's action. OR ii.If the labeling procedures had been carried out on an intact intestinal
epithelial cell, briefly describe how the labeling pattern would compare
(and contrast) with that obtained for erythrocytes. |
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