## Molecular Cell Biology

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### Molecular Cell Biology

Can anyone help me with the answers to (b) and (c)?? I'm completely stuck

Problem 2 - Endocytosis of EGF and a fluid phase-marker (HRP) (70 marks)

Cells take up extracellular molecules by receptor-mediated endocytosis (RME) and by fluid

phase endocytosis (FPE). The efficiencies of these 2 pathways were compared experimentally by

incubation of the human A431 epithelial carcinoma cell line with epidermal growth factor (EGF)

to measure RME, and horseradish peroxidase (HRP) to measure FPE. A quantitative analysis

was carried out by incubating A431 cells with 40 nM ferritin-labelled EGF and 20 μM HRP

(which is at a 500-fold higher concentration than that for EGF) for various times. The cells were

then chemically fixed (cross-linked), stained for HRP-activity and the presence of the electrondense ferritin stain, and examined using the electron microscope. Both EGF and HRP were

present in small vesicles with an internal radius of 20 nm but EGF was very commonly detected

whereas the HRP was rarely detected.

The rates of EGF and HRP uptake were compared in Figure 1. HRP uptake was linear with

respect to both time and concentration: A431 cells took up HRP at a rate of 1 pmol/hr at 20 μM

HRP (Fig. 1A). EGF uptake showed an initial linear phase but reached a plateau at higher EGF

concentrations in the extracellular medium. Using appropriate formulas, answer the following:

(a) Explain why the shapes of the lines in the Fig. 1 are different for HRP and EGF. (10 marks)

(b) Calculate the number of EGF receptors on the surface of each A431 cell. (20 marks)

(c) Calculate how many HRP molecules get taken up into a single endocytic vesicle (radius, r =

20 nm) when the medium contains 1 mg/ml HRP (molecular weight of HRP = 40 000 daltons).

Note: The volume of a sphere = 4/3πr

3

(25 marks)

Problem 2 - Endocytosis of EGF and a fluid phase-marker (HRP) (70 marks)

Cells take up extracellular molecules by receptor-mediated endocytosis (RME) and by fluid

phase endocytosis (FPE). The efficiencies of these 2 pathways were compared experimentally by

incubation of the human A431 epithelial carcinoma cell line with epidermal growth factor (EGF)

to measure RME, and horseradish peroxidase (HRP) to measure FPE. A quantitative analysis

was carried out by incubating A431 cells with 40 nM ferritin-labelled EGF and 20 μM HRP

(which is at a 500-fold higher concentration than that for EGF) for various times. The cells were

then chemically fixed (cross-linked), stained for HRP-activity and the presence of the electrondense ferritin stain, and examined using the electron microscope. Both EGF and HRP were

present in small vesicles with an internal radius of 20 nm but EGF was very commonly detected

whereas the HRP was rarely detected.

The rates of EGF and HRP uptake were compared in Figure 1. HRP uptake was linear with

respect to both time and concentration: A431 cells took up HRP at a rate of 1 pmol/hr at 20 μM

HRP (Fig. 1A). EGF uptake showed an initial linear phase but reached a plateau at higher EGF

concentrations in the extracellular medium. Using appropriate formulas, answer the following:

(a) Explain why the shapes of the lines in the Fig. 1 are different for HRP and EGF. (10 marks)

(b) Calculate the number of EGF receptors on the surface of each A431 cell. (20 marks)

(c) Calculate how many HRP molecules get taken up into a single endocytic vesicle (radius, r =

20 nm) when the medium contains 1 mg/ml HRP (molecular weight of HRP = 40 000 daltons).

Note: The volume of a sphere = 4/3πr

3

(25 marks)

b) for this you need the speed of uptake

c) if you know radius and formula for volume, you should be able to calculate volume, shouldn't you? If you know concentration and volume, you should be able to calculate the weight, shouldn't you? If you know the weight and molecule weight, you should be able to calculate the number of molecules, shouldn't you?

c) if you know radius and formula for volume, you should be able to calculate volume, shouldn't you? If you know concentration and volume, you should be able to calculate the weight, shouldn't you? If you know the weight and molecule weight, you should be able to calculate the number of molecules, shouldn't you?

http://www.biolib.cz/en/main/

*Cis*or*trans*? That's what matters.### Re: Molecular Cell Biology

b) the speed of uptake is 4x106 pmol/hr

where do i go from here?

Thank you for your help

where do i go from here?

Thank you for your help

check out this cool animation on this topic!

http://www.youtube.com/watch?v=he_Qw4_v ... re=related

http://www.youtube.com/watch?v=he_Qw4_v ... re=related

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