Answered: a. You selectively label phospholipids…

Biochemistry

9th Edition

ISBN:9781319114671

Author:Lubert Stryer, Jeremy M. Berg, John L. Tymoczko, Gregory J. Gatto Jr.

Publisher:Lubert Stryer, Jeremy M. Berg, John L. Tymoczko, Gregory J. Gatto Jr.

Chapter1: Biochemistry: An Evolving Science

Section: Chapter Questions

Problem 1P

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Question

a. You selectively label phospholipids with a fluorescent dye and perform the FRAP assay. You detect a single fluorescence molecule move 100 micrometre into the photobleached area in 2.5 seconds. What is the diffusivity constant for this phospholipid? b. Increasing the ratio of unsaturated to saturated phospholipids in a biological membrane increases membrane fluidity. Describe how the FRAP photobleaching and recovery curves would differ between membranes with 63% and 41% unsaturated phospholipids. c. Draw the FRAP photobleaching and recovery curves for your answer in b. d. What does the “immobile fraction” in a FRAP experiment suggest (see diagram)?

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Photobleach
Immobile fraction
1/2
Mobile fraction
Fo
Time
Photobleach
Recovery
3) Fluorescence recovery after
photobleaching (FRAP) is a
microscopy-based technique that can
be used to assess how fluorescently-
labeled proteins and lipids move within
cellular membranes. A specified area of
the membrane is photobleached and
fluorescence signal in that region is
diminished. The same area is then
monitored for recovery of fluorescence
intensity. See diagrams below.
Fluorescence intensity

Expert Solution

Step 1

FRAP is a method to determine the diffusion or kinetics of the biomolecule through tissue or cells. Through this method we can quantify the two dimensional lateral diffusion of fluorescently labeled biomolecules. It is a significant method to understand the diffusion of molecules through the cell membrane or binding or interaction of protein.

It is also used to study the mobility of individual lipid molecules within a cell membrane.