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Unformatted text preview: Biol110 Cell Biology Problem Set 5 Key Michael F. Rexach, Associate Professor, MCD Biology E-mail: [email protected] Question #1: How would you measure directly whether a particular membrane or lumenal cargo protein is actively enriched in vesicles that are budding? Do you expect a very abundant soluble protein to concentrate in vesicles via receptors as vesicles bud? How about a protein that is very rare in abundance? Explain. The best way to physically measure a protein’s enrichment in vesicles (using cells or in reconstituted vesicle budding reactions) is to use quantitative immunoelectron microscopy. You need an antibody against the protein you wish to analyze. The antibody would be coupled to 10 nm gold particles and you should perform an immuno-EM experiment. You have to compare the “density” or enrichment of IgG-gold particles within a vesicle bud in comparison to the distribution of the gold particles throughout the lumen or membrane of the ER. Enriched proteins will appear concentrated in vesicle areas in comparison to the distribution of the protein in the ER lumen. Very abundant proteins that need to be secreted probably do not accumulate in the vesicles actively; they probably exit the ER by “bulk flow.” It would be impractical to further concentrate abundant proteins; and would also be impractical to have abundant receptors for them. Most average proteins likely concentrate in budding vesicles using receptors that tether themselves to the vesicle coat proteins, and thus exit the ER by recruitment into vesicles. recruitment into vesicles....
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This note was uploaded on 06/23/2008 for the course BIO 110 taught by Professor Rexach during the Spring '08 term at UCSC.
- Spring '08
- cell biology