Lecture 6: Membrane Structure

Question 1

What factors play a role in membrane fluidity

Answer 1

-lipids and cholesterol
- proteins for movement across the membrane

Question 2

What recent modifications have been made to the fluid mosaic model

Answer 2

the movement of lipid and membrane proteins differs significantly in pure lipds compared to those in biological membranes

Question 3

How can the mobility of lipids and membrane proteins be measured

Answer 3

FRAP

Question 4

Describe FRAP stepwise

Answer 4

Fluorescence Recovery After Photobleaching:
1) Label cell surface molecule
2) measure the starting intensity of fluorescence
3) Use a high-intensity laser to bleach the fluorophore in a small part of the membrane
4)Measure the rate of fluorescence over time to determine how quickly the level of fluorescence is recovered

Question 5

What does bleaching refer to in the FRAP process

Answer 5

Using a high-intensity laser to damage the fluorescent molecule on the surface of the protein
** does not affect the mobility of the protein, it can still move just cannot be detected**

Question 6

What does a high rate of fluorescence recovery in FRAP suggest

Answer 6

Molecule is very mobile within the membrane, can move quickly

Question 7

Why might some proteins or ligands spend more time interacting within certain regions of the membrane

Answer 7

eg lipid rafts
may have functional roles

Question 8

What is the lateral diffusion of proteins dependant on

Answer 8

Interactions with other proteins such as the cytoskeleton or with extracellular components for signalling and transport

Question 9

Why can a strong detergent like SDS not be used when preforming a FRAP experiment

Answer 9

proteins will denature, providing no results

Question 10

What kind of PTMs are preformed proteins and lipids that help them enter the membrane

Answer 10

Addition of sugars to lipids and proteins mediated by enzymes is important for membrane insertion and cell recognition

Question 11

How does membrane transport necessitate an asymmetric membrane

Answer 11

Exterior pf membrane contains transport and signalling molecules such as:
- oligosaccharide chains of glycoproteins
- lipid-anchored proteins
-peripheral membrane proteins
Cytosol side of membrane contains other molecules, such as integral membrane proteins

Question 12

Describe uncatalyzed membrane diffusion

Answer 12

uncatalyzed transbilayer ‘flipflop’ diffusion occurs when a phospholipid just flips over to the other side, requires hydrophyllic head to travel through large hydrophobic section

Very slow, takes days

Question 13

What are three enzymes that catalyze the process of membrane diffusion

Answer 13

• flippase
• floppase
• scramblase

Question 14

What is the difference between flippase and floppase

Answer 14

flippase is a P-type ATPase and moves PE and PS (specifically) from exterior to cytosolic side

floppase is an ABC transporter (still needs ATP) moves phospholipids from the inner cytosolic side to the exterior

Question 15

Describe scramblase

Answer 15

An ATP-independant enzyme that moves lipids in either direction; movement down the concentration gradient
- eliminates any asymmetry due to lipid distribution

Question 16

What do enzymes catalyze on the extracellular surface of the membrane

Answer 16

Catalyze the addition of oligosaccharides to proteins and lipids

Question 17

Define lipids

Answer 17

class of molecules involved in providing structural support for cells and organelles, storage of carbons for energy, and that can play a role in information transduction and signalling

Question 18

What are the physical properties of lipids

Answer 18

low solubility in water, high solubility in non-polar solvents

Question 19

What are the four possible structures of lipid “membranes”

Answer 19

monolayers, bilayers, miscelles (inside-out and normal), unilamellar vesicle (liposome; most similar to cell membrane)

Question 20

What 4 types of lipids are used for structural support

Answer 20

• phospholipids
• sphingolipids
• glycolipids
• sterols

Question 21

What type of lipids are used for signalling

Answer 21

• eicosanoids
• sterols

Question 22

How can double bonds in fatty acids be numbered

Answer 22

typically numbered relative to the carboxyllic acid (delta#), but can also be numbered from the terminal carbon (omega)

Question 23

What type of bond is used to join fatty acids to a head group

Answer 23

ester bonds

Question 24

How is the carbon skeleton of a fatty acid named

Answer 24

carbons : # or double bonds
monounsaturated; one double bond
polyunsaturated (PUFA): more than one double bond

Question 25

What are two polyunsaturated essential fatty acids for humans

Answer 25

omega-3 and omega-6 fatty acids are PUFAs that humans cannot synthesize and must be obtained form our diet (thus essential)

Question 26

How does membrane fluidity affect melting temperature

Answer 26

low membrane fluidity, like in saturated fatty acids results in high melting temperature because more energy is required to break the non-covalent interactions high membrane fluidity, like in unsaturated fatty acids or a mix of both results in a low melting point

Question 27

What phase transition characterizes the melting of membrane lipids

Answer 27

phase transition from a gel-like solid to the liquid crystalline phase

Question 28

What distinguished pure liquid samples from native membranes on phase transition diagrams

Answer 28

pure liquid samples have sharp, well-defined transition temperatures while native membranes have broad peaks

Question 29

What are TAGs? Describe their structure and function

Answer 29

Tri-acyl-glycerols are a dehydrated storage form of lipids found primarily in adipocyte cells that are an efficient, unlimited energy reserve of reduced carbon chains Structure is three fatty acids bound to a glycerol backbone through ester linkages; fatty acid tails may contain double bonds (mixed triglycerides)

Question 30

What are amphipathic molecules, how are they made, and what are the two types

Answer 30

Amphipathic molecules contain both polar and nonpolar groups made by attaching fatty acid chains to a polar head group (OH, sugar, phosphate) Two types are glycero-phospho-lipids and sphingolipids

Question 31

What is the most common steroid found in membranes

Answer 31

cholesterol

Question 32

What are two examples of glycerophospholipids

Answer 32

phosphatidylethanolamine (PE) PS (-serine, positive charge)

Question 33

What are two examples of sphingosines

Answer 33

sphingomyelin gangliosides both have sugar head groups

Question 34

name two functions of cholesterol

Answer 34

1) metabolized to other other hormones (cortisol, testosterone) and bile salts- needed for dietary lipid absorption 2) Form lipid rafts to help moderate membrane fluidity and play a role in signal transduction

Question 35

how are lipid rafts formed

Answer 35

cholesterol forms complexes with sphingolipids, glycolipids, and some lipid-anchored proteins

Question 36

What type of membrane has the highest percentage cholesterol? which has the least?

Answer 36

most: plasma least: inner mitochondrial

Question 37

Describe peripheral membrane proteins and removal conditions

Answer 37

adhere to the surface of lipid membranes or integral membrane proteins through non-covalent interactions and can be removed using mild conditions such as a pH change or increase in salt

Question 38

Describe integral membrane proteins and removal conditions

Answer 38

completly span the membrane (20AAs) and require harsh conditions such as detergents or organic solvents to be removed (purified)

Question 39

Describe lipid-anchored proteins, removal conditions, and name a subtype

Answer 39

Lipid chains covalently attached to amino acid functional groups and side chains Require harsh detergents or solvents for membrane removal GPI-anchored proteins (glycosyl phosphatidylinositol-) are covalently linked through a sugar chain to a lipid anchor

Question 40

How do detergents solubilize membrane proteins

Answer 40

detergents are amphipathic molecules that create micelles around hydrophobic regions to solubilize membrane proteins so they can then be purified and analyzed

Question 41

What is the critical micelle concentration

Answer 41

concentration at which the detergent spontaneously forms stable micelle structures

Question 42

How many amino acids are needed to span the membrane in the ALPHA HELIX structure

Answer 42

20

Question 43

how do you read the Kyte-Doolittle Hydropathy scale

Answer 43

Each residue has a given hydropathy index. Positive values are more hydrophobic and therefore inside the membrane. Negative values are hydrophillic and polar, and therefore outside the membrane

Question 44

how can the hydropathy index for a stretch of amino acids be determined

Answer 44

By averaging the kyte-doolittle hydrophobicity values of all the amino acids found in the segment average of 7 values gets assigned to the middle amino acid, window shifts one to the right to recalculate and average is assigned to the next AA and so on

Question 45

How can the window size in calculating hydropathy index be changed

Answer 45

window can be 7, 9, 11, or 13 Smaller window gives noisier plots larger window gives smoother plots

Question 46

How can transmembrane segments be identified when reading a hydropathy index

Answer 46

Identify a broad peak over zero that spans over 20 amino acids

Question 47

What are two examples of membranes containing transmembrane segments

Answer 47

Glycophorin A (GPA) contains a single transmembrane segment between AA 72-91 Bacteriorhodopsin contains 7 transmembrane segments, or positive peaks that span at least 20 AAs