Cell membranes

• • Polar phosphates face the membrane surfaces
  • Nonpolar fatty acid tails face into the interior of the membrane

  Lipid bilayer prevents random movement of substance in and out of the cell
• Fatty acids

  • Fatty acids are long, unbranched hydrocarbons chains
  • Fatty acids can be saturated or unsaturated
  • Naturally occurring fatty acids have cis double bonds
• Phosphoglycerides:

  • built on glycerol backbone
  • includes: glycerol + 2 fatty acid chains + phosphate group + additional group
  • Often one unsaturated and one saturated fatty acid chain
  • additional group determines the identity
• Label head groups charge using symbols except for neutral

  A) neutral

  B) -

  C) neutral

  D) -

  E) -

  
• Sphingolipids:

  • built on sphingosine
  • longer and more saturated fatty acid chains
  • amphipathic (both hydrophobic and hydrophillic regions)
  • Additional groups can be added to head group
  • roles in signal transduction, membrane structure, sensins

  Most basic sphingolipid = ceremides: sphingosine + fatty acid

  Sphingomyelin is a sphingolipid that is also a phospholipid
• If additional group to sphingolipid is a carbohydrate, the molecule is a glycolipid
• Cholesterol:

  • amphipathic
  • Oriented with small hydrophobic group facing the membrane surface
  • Remainder is embedded in the fatty acid tails of phospholipids
  • They impair the movement b/c of their rigidity (due to their rings) of the fatty acid tails of phospholipids
• Practice !

  1. phosphate, additional group attached to phosphate, glycerol, fatty acid chains

  3. a, b, c, and d

  A) Phospholipids

  B) sphingolipids

  C) phosphoglycerides

  
• Membrane asymmetry:

  Asymmetry affects membrane permeability, surface charge, membrane shape and stability.

  Examples:

  • PE: Promotes curvature of the membrane (because it has the smallest additional groups)
  • PS: Negative charge interacts with transmembrane proteins
  • PI: Roles in signal transduction
• Cholesterol is relatively symmetrical between the two sides of the membrane
• All membrane carbohydrates in the plasma membrane face the outside of the cell
• Lipid composition can determine:

  • The physical state of the membrane
  • Facilitate protein interactions
  • Roles in signal transduction
• Major membrane functions:

  1. compartmentalization
  2. Scaffold for biochemical activities (proteins can be structured within a membrane to facilitate interactions)
  3. Selectively permeable membrane
  4. Solute transport
  5. Response to external stimuli
  6. Cell-cell communication
  7. Energy transduction ( concentration gradients between the two sides)
• Fluidity is influenced by temperature (among other things)

  • transition temperature (TT) or melting temperature is the intersection between the crystalline gel phase (below TT) and the liquid crystalline phase (above TT)
• Transition temperature (and thus fluidity) is affected by:

  1. Fatty acid chain saturation:
  2. Saturated= stronger van der waals interactions (more rigid)
  3. Unsaturated = kinks at double bonds, impairing the interactions (more fluid)
  5. 2. Cholesterol content:
  6. Flat, rigid, hydrophobic rings impair the movement of the fatty acid tails
  7. Thus, eliminating a sharp transition temperature and creating intermediate fluidity.
  8. 3. Fatty acid chain length:
  9. - shorter fatty acid tails = less interactions (more fluid)
• Maintaining the ideal membrane fluidity is important for:

  • maintaining structural organization and mechanical support
  • Enabling interactions (clustering of proteins)
  • Membrane assembly/cell growth/ cell division
  • Cell movement, secretion, and endocytosis
• How can cells alter the lipid composition of the membrane?

  In response to colder temps:

  • decrease fatty acid tail length
  • desaturate double bonds (enzyme called desaturase can do just that)
  • Reshuffle fatty acid chains between phospholipids so there is more phospholipids with two unsaturated chains (greatly lowers melting temp)

  The opposite would be true for higher temps
• Does increasing the % of saturated fatty acids increase or decrease melting temp?

  Increases melting temp because the membrane is more rigid because of more van der waals interactions (ie, you need a higher temp (more energy) to transition between crystalline gel to crystalline fluid)
• Membrane proteins are distributed asymmetrically across the two leaflets of the membrane bilayer
• Integral membrane proteins:

  • permanently anchored or are part of the membrane

  A) monotopic

  B) bitopic

  C) polytopic

  
• transmembrane proteins pass through the lipid bilayer and contain one or more transmembrane domain(s). Part of integral membrane protein class.

  • act as receptors, channels, or have roles in electron transport
  • transmembrane domains tend to be hydrophobic and interact with the fatty acids of the membrane (string of ⁓ 20 non-polar amino acids)
  • at the surface of the membrane, protein is hydrophilic
• Peripheral membrane proteins:

  • associated to the membrane through weak non-covalent bonds
  • dynamic: can be recruited/released from the membrane
  • roles in: signal transduction, mechanical support for membrane, anchor for integral proteins, enzymes
  • Mostly hydrophilic/polar since they are interacting withing the aqueous environment
• Lipid-anchored proteins:

  • either on the extracellular or cytoplasmic side
  • Covalently-linked to lipid molecule within the bilayer

  1. GPI-anchored proteins: proteins attached to the membrane by a small oligosaccharide complex linked to phosphatidylinositol (PI) in the membrane.
  2. Face the extracellular space and have roles in cell adhesion and as receptors
  3. 2. Hydrocarbon chains embedded in the lipid bilayer: directly linked to a lipid embedded into the membrane (as opposed to a complex)

  • Found in the cytoplasmic leaflet
  • roles in signal transduction

  A) GPI-anchored

  B) Lipid-anchored

  
• Phospholipid dynamics:

  • phospholipids can easily move laterally across the same leaflet
  • phospholipids cannot flip-flop between the two leaflets easily (transverse diffusion)
  • enzyme flippase can help phospholipids flip and establish membrane asymmetry
• Membrane protein dynamics:

  1. random diffusion
  2. immobilized (no movement)
  3. Particular direction (motor proteins)
  4. restricted by other integral membrane proteins
  5. restricted by membrane skeleton proteins
  6. restrained by extracellular material (tangled up in stuff)

  A) 5

  B) 3

  C) 4

  D) 6

  E) 2

  F) 1

  
• Secondary active transport: using concentration gradients to move molecules across the membrane.


  • one substance with its concentration gradient to provide energy for the other moves against

  Symporter/cotransporter: transports two substances in the same direction.

  Antiporter/exchanger: transports two substance in opposite directions
• Na+/glucose cotransporter

  • transports glucose from the intestinal lumen into epithlial cells
  • Na+ ions concentration is low inside cell

  Na+ moves into the cell with its concentration gradient which is used to drive glucose into the cell
• FRAP (Fluorescence recovery after photobleaching)


  Can be used to study the membrane dynamics in vivo

  1. membrane is tagged with fluorescent dye
  2. portion of membrane is photobleached
  3. The rate of recovery of fluorescence is a measure of the rate of diffusion (fluidity of membrane)
• Isolating peripheral membrane proteins

  1. lysing the cells and separating the plasma membrane
  2. pellet 1: insoluble material- membrane
  3. supernatant 1: soluble material
  4. 2. Salting out: salt will compete with peripheral membrane proteins and disrupt interactions with the membrane.
  5. peripheral membrane proteins in supernatant 2
• Isolating transmembrane proteins

  1. lysing the cells and separating the plasma membrane
  2. pellet 1: insoluble material- membrane
  3. supernatant 1: soluble material
  4. 2. Salting out: salt will compete with peripheral membrane proteins and disrupt interactions with the membrane.
  5. peripheral membrane proteins in supernatant 2
  6. 3. add strong detergent: detergent is amphipathic and can substitute for phospholipids to stabililze transmembrane proteins and make them soluble
  7. transmembrane protein in supernatant 3
• Isolating GPI-anchored proteins

  1. 3. add strong detergent: detergent is amphipathic and can substitute for phospholipids to stabililze transmembrane proteins and make them soluble
  2. transmembrane protein in supernatant 3
  3. 4. GPI-anchored proteins are linked to phosphatidylinositol in the membrane. Found in detergent-resistent portions of the membrane.

  • Need to treat with phosphatidylinositol-specific phospholipase C (PI-PLC) which cleaves the protein from the membrane
  • GPI-anchored protein found in supernatant 4
• SDS-PAGE

  Seperates proteins by size

  • SDS is a negatively charged amphipathic detergent. It denatures the protein and coats the protein with a negative charge
  • negatively-coated proteins migrate towards the positive end with smaller proteins moving further down.
• Glycolipids

  • If a simple sugar is added, then the glycolipid is a cerebroside.
  • If a cluster of sugar that includes sialic acid is added to the sphingolipid, then the glycolipid is a ganglioside