Lecture 7

Question 1

Fluid Mosaic Model

Answer 1

plasma membrane is a collection of several types of molecules (phospholipids, cholesterols, and proteins) that are constantly moving

Question 2

Integral (transmembrane protein)

Answer 2

embedded in lipid bilayer

Question 3

Peripheral protein

Answer 3

inner or outer surface of lipid bilayer ; attached to integral protein through noncovalent interactions

Question 4

How do integral (transmembrane proteins) need to be analyzed in experiments?

Answer 4

need to be extracted with detergents (molecules that disrupt cell membranes)

Question 5

How do peripheral proteins need to be analyzed in experiments?

Answer 5

can be removed with high salt

Question 6

What is the purpose of detergents?

Answer 6

Disrupt cell membranes

Question 7

Ionic detergents (SDS)

Answer 7

have a charged head group; they will disrupt membranes and denature / unfold proteins

Question 8

Non-Ionic detergents (Triton X-100)

Answer 8

have a polar uncharged head group ; disrupt membranes but will not denature most proteins and can denature partially unfolded proteins

Question 9

2 steps to analyze transmembrane proteins with biochemical techniques:

Answer 9

1. Expose whole cell lysate to strong detergents to generate water soluble membrane proteins
2. Analyze solubilized membrane proteins using immunoprecipitation / Coomassie / Western blot

Question 10

Strategies to restrict movement of membrane proteins

Answer 10

1. Barriers can restrict membrane proteins to a particular region of the cell
2. Membrane proteins can be tethered to the extracellular matrix (ECM) outside cell and cytoskeleton inside cell
   - Use of Inegrins and Cadherins

Question 11

Inegrins (membrane proteins)

Answer 11

bind to ECM (extracellular matrix) and cytoskeleton; helps anchor cells to ECM to build tissues

Question 12

Cadherins (membrane proteins)

Answer 12

cell adhesion molecules that allow cells to adhere to each other

Question 13

Fluorescence Recovery After Photobleaching (FRAP)

Answer 13

technique used to analyze mobility of lipids and proteins in plasma membrane

Question 14

Steps on how FRAP is performed

Answer 14

1. Membrane proteins or lipids are labeled with fluorescence
2. Lazer irreversibly bleaches fluorescent dye in specific area (bleached area)
3. Cell monitored with fluorescence microscope to assess how much time it takes for neighboring unbleached proteins/ lipids to migrate into bleached area
4. Rate of fluorescence recovery is used to assess fluidity of membrane

Question 15

Slope of bleach increases (time vs. fluorescence in bleached area) after exposed to bleach from FRAP experiment indicates

Answer 15

Protein is mobile, fluorescence recovers quickly in bleached area

Question 16

Slope of bleach does not increase after exposed to bleach (time vs. fluorescence in bleached area) from FRAP experiment indicates

Answer 16

Protein is not mobile, fluorescence does not recover in bleached area

Question 17

Steps for Cell Fusion to study movement of membrane proteins.

Answer 17

1. Different fluorescent labels were attached to membrane proteins on mouse cells and human cells
2. Labeled cells were fused to create hybrid cell
3. Hybrid cell was imaged 40 minutes after cell fusion with fluorescence microscope