Lipids, Membranes and Transport Outline
Transcript of Lipids, Membranes and Transport Outline
-
7/29/2019 Lipids, Membranes and Transport Outline
1/25
Lipids, Membranes and Transport Outline
1 | P a g e
Biological Membraneso Framework of the membrane is the phospholipid bilayero Phospholipids are amphipathic (forms bilayer) molecules
Hydrophobic (water-fearing) faces in Hydrophilic (water-loving) faces out
o Membranes contain Proteins and carbohydrates Fluid Mosaic Model
o The membrane is considered a mosaic of lipid, protein and carbohydrate moleculeso Membrane resembles a fluid- lipids and proteins move around relative to each other
Lateral movement across membrane Membrane Proteins
o Integral or Intrinsic membrane proteins Transmembrane Proteins
Region physically embedded in the hydrophobic region of bilayer Lipid anchored Protein
Noncovalent attachment of a lipid to amino acid side chain within a proteino Peripheral membrane (extrinsic proteins)
Noncovalently bound to regions of integral membrane proteins that project out frommembrane or bound to polar head groups of phospholipid
o Hydrophilic Head- loves watero Hydrophobic Tails- hates water
Hydrophobic
Amino acids
-
7/29/2019 Lipids, Membranes and Transport Outline
2/25
Lipids, Membranes and Transport Outline
2 | P a g e
Approximately 25% of all genes Encode membrane proteinso Membranes are extremely important- biologically and medically
Membranes are Semifluido Fluidity
Individual molecules remain in close association but have ability to move within themembrane
o Semifluid Most lipids can rotate freely around their long axes and move laterally within the
membrane leaflet
Flipflop of lipids from one leaflet to the opposite leaflet does not occur spontaneously Flippase requires ATP to transport lipids from one leaflet to another
2. Multiple
alpha helices
1.Alpha-helix
3. series of
alpha helices4. 1 layer
of bilayer
5. Lipid
anchored
-
7/29/2019 Lipids, Membranes and Transport Outline
3/25
Lipids, Membranes and Transport Outline
3 | P a g e
Factors affecting Fluidityo Length of fatty acyl tails
Hydrophobic tails- the shorter the tail, the more fluid it iso Presence of double bonds in the acyl tails
Big factor in fluidity Double bond creates a kink in the tail, making it more difficult to interact with
nearby tails and making the bilayer more fluido Presence of Cholesterol
Tends to stabilize membranes Depends on temperature
Cold-rigid Warm-fluid
Saturationo Saturated is when there are no C=C double bondso Unsaturated is when there are C=C double bonds
Cholesterol Stabilizes Membraneso High Temperatures
Lipid becomes fluid Cholesterol becomes more rigid
o Low Temperatures Lipid becomes more rigid Cholesterol becomes more fluid
Loves water
Hates water
Head
Tails
Head
2 C=C
bonds
1 C=C
bonds
1 C=C
bonds
0 C=C bo
Saturated
-
7/29/2019 Lipids, Membranes and Transport Outline
4/25
Lipids, Membranes and Transport Outline
4 | P a g e
Lateral TransportExperimento Mouse and Human cells fused togethero Added a florescent label antibody and sawo 0 C- Cold temperature
Protein unable to move laterally and remains on one side of the cello 37 C- Higher temp
Movement observed, the protein is on both sides of the cell Some Integral Membrane Proteins have restricted movement
o 10%-70% of membrane proteins may be restricted in their movement Anchored, attached to cytoskeleton or extracellular matrix
o Membrane proteins may be attached to molecules outside of the cell, so the network of proteinsthat forms the extracellular matrix
How to study Membraneso TEM (Transmission Electron Microscopy)
Thin section sample stained with heavy metal dyes, gives electron dense, binds topolar bits since metal has a charge
Train track looking membrane bilayer (polar heads dyed)
Transmembrane
Protein
-
7/29/2019 Lipids, Membranes and Transport Outline
5/25
Lipids, Membranes and Transport Outline
5 | P a g e
o FFEM (Freeze Fracture Electron Microscopy Specialized form of TEM, can be used to analyze the interiors of phospholipid bilayers Sample is frozen in liquid Nitrogen and fractured with a knife Leaflets separate into P face (protoplasmic) and E face (extracellular) Provide 3D detail about membrane protein form and shape Using FFEM- Branton in 1966 was able to conclude that membranes are formed from
phospholipid bilayers, with proteins intercalated within
Synthesis of Membrane Componentso In Eukaryotes, Cytosol and endomembrane system work together to synthesize most lipidso Process occurs at cytosolic leaflet of smooth ERo Fatty acid building blocks made via enzymes in cytosol or taken into cells from food
-
7/29/2019 Lipids, Membranes and Transport Outline
6/25
Lipids, Membranes and Transport Outline
6 | P a g e
All phospholipids are synthesized in the Smooth ER Transfer of Lipids to other Membranes
o Lipids in ER membrane can diffuse laterally to nuclear envelopeo
Transported via vesicles to Golgi, lysosomes, vacuoles, or plasma membrane (blebbing)o Lipid exchange proteins- extract lipid from one membrane for insertion in anothero 2 Phospholipids into 1
Membrane Protein Synthesiso Most transmembrane proteins are directed to ER membraneo From the ER, membrane proteins can be transferred via vesicles to other regions of the cello Trans-membrane (expand layer) proteins are translated directly into the phospholipid bilayer
of the rough ER
Stretches of 20 hydrophobic amino acids (universal for transmembrane proteins) Tends to form alpha-helical structure Length of region is 20 amino acids long
How transmembrane proteins are translated
Protein traffikingo Proteins destined for any part of the endomembrane system or secretion from the cell are
translated directly into the ER
Anything membrane bound is directly translated into the ER membrane itself- evensome non-membrane proteins also
Protein finished
being translated
-
7/29/2019 Lipids, Membranes and Transport Outline
7/25
Lipids, Membranes and Transport Outline
7 | P a g e
o How proteins are sorted once in the endomembrane system is reliant on the physical propertiesof lipid vesicles
- Like a lava lamp- things can bud off and go back and forth, not unidirectional
-Most things can go back and forth with vesicles, not unidirectional
-
7/29/2019 Lipids, Membranes and Transport Outline
8/25
Lipids, Membranes and Transport Outline
8 | P a g e
Protein Sorting to the ER
Protein Sorting to the Golgio From ER to Golgio Uses membrane proteins V-Snare and T-Snare to merge protein-containing vesicles to the correct
target
o Certain proteins are classified as V-Snare or T-Snare
Signal
recognition
protein
(SRP)
Docks with
protein in
Rough ERto translate
Translated
into ER
lumen
Transmembrane
Protein- 20 amino
acids, forms alpha
helice
Polypeptide
chain- inside
ER lumen
Proteins that
form matrix
(cage)
Pinches off
(Blebs off)
Attaches to
T-Snare
Gol i
Rough ER
Inside ER Lumen
-
7/29/2019 Lipids, Membranes and Transport Outline
9/25
Lipids, Membranes and Transport Outline
9 | P a g e
Protein Sorting in the Golgi (continued)o Cargo protein, targeted to the golgi, binds to a cargo receptor protein in the ER membrane. Coat
proteins on the outside aid in the formation of vesicle buds, which will contain the appropriate
cargo. In the vesicle membranes are also V-snares.proteins embedded in the membrane which
target the vesicle to the correct site. Upon approaching the target (Golgi), T-snare proteins
within the golgi membrane bind with the v-snares, causing the membranes to fuse, and the cargo
protein to be released into the golgi.
Coat Proteins cause the membrane to Bleb Off
SARE Proteins allow vesicles to join together- (Direct mechanism)o 2 Nerves
- Transmembrane Proteins
-
7/29/2019 Lipids, Membranes and Transport Outline
10/25
Lipids, Membranes and Transport Outline
10 | P a g e
Glycosylationo Process of covalently attaching a carbohydrate to a protein or lipid
Glycolipid- Carbohydrate to lipid (sugar stuck to fats) Glycoprotein- Carbohydrate to protein (sugar stuck to protein)
o Proteins ER to Golgio Can serve as recognition signals for other cellular proteinso Plays a role in cell surface recognitiono Protective effects
Cell coat or glycocalyx- carbohydrate rich zone on the cell surface shielding cell
Clicker Questiono The bond that attaches Carbohydrates to Lipids and Proteins in the Plasma Membrane
Covalent Bond Because were transferring to O or N
Spontaneously Happens
-Forms one larger unit
-T-Snare attaching to Golgi
-
7/29/2019 Lipids, Membranes and Transport Outline
11/25
Lipids, Membranes and Transport Outline
11 | P a g e
Protein Glycosylationo N-Linked
In ER & Golgi Attachment of Carbohydrate (sugars) to Nitrogen (N) atom of asparagine side chain
o O-Linked Only in Golgi Addition of sugars to Oxygen (O) atom of Serine or Threonine side chains
Both have hydroxyl groups (OH), so sugars get added on there N-Linked Glycosylation
o Carbohydrate (sugars) to N atom of asparagine side chaino In ER & Golgio Dolichols are previously made in the ERo Transfer to target Asp sequenceo Commonly found in membrane proteins
-
7/29/2019 Lipids, Membranes and Transport Outline
12/25
Lipids, Membranes and Transport Outline
12 | P a g e
O-Linked Glycosylationo Occurs only in Golgio Links sugar chains to the O in Ser or Thr side chainso Important for production of proteoglycans
Mucous Extracellular matrix
o Transfers sugar group onto OH of Ser and Thr Endocytosis and Exocytosis
o Transport larger molecules such as proteins and polysaccharides, and even very large particleso Usually larger things gets endocytosed or exocytosed
Ex. Load of Insulin excreted from Pancreatic cello Exocytosis
From inside cell to outside cell Material inside the cell packed into vesicles and excreted into the extracellular matrix
o Endocytosis Plasma protein invaginates or folds inward to form a vesicle that brings substances into
the cell
Receptor-mediated endocytosis Pinocytosis- Small molecules in extracellular environment are brought into the cell
Used for absorption of extracellular fluids Phagocytosis-Eat Endocytose bacteria
Engulfing solid particles by the cell membrane Defense mechanism- to remove cellular debris- immune response
Exocytosiso Inside cell to Outside the cell
-
7/29/2019 Lipids, Membranes and Transport Outline
13/25
Lipids, Membranes and Transport Outline
13 | P a g e
Endocytosiso Into the Cell, receptor-mediatedo Blebs off into the cello Guided by cytoskeleton (carries cargo)
Phospholipid Bilayer Barriero Serves as a barrier to hydrophilic molecules and ions due to hydrophobic interior and hydrophilic
outside
o Rate of diffusion depends on chemistry of solute and its concentrationo Gasses and a few uncharged molecules can passively diffuse acrosso Ions and large polar molecules diffuse slowly
- Things that can get thru freely- Hydrophobic
molecules. Gasses, small molecules
-This is how cells breathe Diffusion
-Diffuse thru membrane freely
Small polar molecules diffuse thru
somewhat slow, uncharged.
Large molecules, more charged andharder to diffuse, cant go thru bilayer
-
7/29/2019 Lipids, Membranes and Transport Outline
14/25
Lipids, Membranes and Transport Outline
14 | P a g e
Membrane Transporto Selectively permeable plasma membrane
Outside environment cant dictate inside environmento Structure ensures
Essential molecules enter Metabolic intermediates remain Waste products exist
Movement Across Membraneo Passive transportdoesnt require an input of energy- down or with gradient
Passive Transport Diffusion of a solute through a membrane without transport protein
Facilitated Diffusion Diffusion of a solute through a membrane with the aid of a transport protein
o Active transport requires energy- up or against gradient
-Simple diffusion
-No transport protein
-Down/with gradient, no
energy required
Ex.) O2, CO2
-Diffusion with Aid oftransport protein
-Down/with gradient, no
energy required
Spontaneous, favorable
G = -
-Up/Against gradient
-Requires energy
Non-spontaneous,
unfavorable
G = (+)
-
7/29/2019 Lipids, Membranes and Transport Outline
15/25
Lipids, Membranes and Transport Outline
15 | P a g e
Active Transporto Movement of a solute across a membrane against its gradient from a region of low to higher
concentration (Low Concentration High Concentration)
o Energetically unfavorable, non-spontaneouso Requires input of Energy to drive movemento Primary Active Transport
Uses a pump- directly uses energy to transport soluteo Secondary Active Transport
Very common Uses pre-existing gradient to drive transport of solute Relying on concentration gradient- ATP is not always used
Cells Maintain Gradientso Living cells maintain a relatively constant internal environment different than outside the cello Transmembrane gradient
Concentration of a solute is higher on one side of a membrane than the othero Ion electrochemical gradient
Both an electrical gradient and chemical gradient
Low Co
High Co
Glucose
Transmembrane Gradient
Conc. Of glucose is higher
outside than it is inside the
cell
-
7/29/2019 Lipids, Membranes and Transport Outline
16/25
Lipids, Membranes and Transport Outline
16 | P a g e
Tonicityo Comparison of solute concentrations across a barriero In relation to the cell
Isotonic- Equal amounts of solute inside and out of the membrane (and equal amounts ofwater)
Hypertonic- Solute concentration is higher on one side of the membrane (and waterconcentration lower)
Hypotonic- Solute concentration is lower on one side of the membrane (and waterconcentration is higher)
Ion Electrochemical
Gradient
-Electrical and Chemical
gradient
-More Na+ outside the cell
and higher Positive charge
outside the cell
-Solute concentration outside the cell is Isotonic to the
inside of the cell
-Solute Concentration outside the cell is Hypertonic to th
inside of the cell
-Solute concentration outside the cell is Hypotonic to the
inside of the cell
-
7/29/2019 Lipids, Membranes and Transport Outline
17/25
Lipids, Membranes and Transport Outline
17 | P a g e
Osmosiso The movement of water towards a soluteo Water diffuses through a membrane from an area with more water to an area with less watero If solutes cannot move, water movement can make the cell shrink or swell as water leaves or
enters the cell
o Osmotic Pressure- The tendency for water to move into any cello Animal cells must maintain a balance between extracellular and intracellular solute
concentrations to maintain their size and shape
o Crenation-shrinking in a hypertonic solution- (can kill cells this way) Aquaporins
o Channel proteins that move water across membraneo Discovery of Aquaporins
CHIP28 Water passively diffuses across plasma membrane Certain cell types allow water to move across the plasma membrane at a much
faster rate than would be predicted by passive diffusion
Peter Agre and colleagues first identified a protein that was abundant in RBCsand kidney cells, but not found in many other cell types
Striking difference was observed between frog oocytes that expressed CHIP28versus the control
CHIP28- renamed Aquaporins Transport Proteins
o Transmembrane proteins that provide a passageway for the movement of ions and hydrophilicmolecules across membranes
o 2 classes bases on movement type Channels- forms a pore to shuttle thru Transporters- Carrier proteins, transforms and changes shape to pass through
-
7/29/2019 Lipids, Membranes and Transport Outline
18/25
Lipids, Membranes and Transport Outline
18 | P a g e
Transporterso Carrierso Conformational change transports soluteo Principal pathway for the uptake of organic molecules
Sugars, amino acids, nucleotides
o Types of Transporterso Uniporter- Single molecule/ion moves in and out of the cell o Symporter/Cotransporter- 2 or more ions or molecules transported in same
direction
o Antiporter- 2 or more ions or molecules transported in opposite directions Glucose Transporters
o Some glucose transporters are uniportero Ex.) Blood brain barrier (BBB), glucose is transported across endothelial cells of small
blood vessels into astrocytes
o Uniporters that transport Glucose down its concentration gradiento Glucose transporters undergo conformational changes that result in a reorientation of
their substrate binding sites across membranes
-
7/29/2019 Lipids, Membranes and Transport Outline
19/25
Lipids, Membranes and Transport Outline
19 | P a g e
Channelso Form an open passageway for the direct diffusion of ions or molecules across the
membrane
o Most are gatedo Ion Channels
Hydration of Ionso Ions in aqueous solutiono Influences their flux through Transmembrane poreso Salts dissolved in water form hydrated ionso The hydrophobicity of lipid bilayers is a barrier to movement of hydrated ions across cell
membranes
Ion channels Dehydrate Ionso All ion channels dehydrate ionso Hydration shell must be removed for ions to be selectively shuttled through a channelo Allow for the rapid selective transport of ions across membraneso Dehydration of Ions costs energy, whereas hydration of ions frees energyo Almost every ion channel is gated
Electrochemical gradientso Across the cell membrane generate the Membrane Potentialo Nernst Equation- used to calculate the membrane potential as a function of ion concentrationso Cells maintain a negative resting membrane potential with the inside of the cell slightly more
negative than the outside
o Membrane potential is a prerequisite for electrical signals and for directed ion movement acrosscellular membranes
-
7/29/2019 Lipids, Membranes and Transport Outline
20/25
Lipids, Membranes and Transport Outline
20 | P a g e
K+ Potassium Channelso Catalyze selective and rapid ion permeationo Function as water-filled pores that catalyze the selective and rapid transport of K+ ionso Complex of 4 identical subunits, each of which contributes to the pore (tetramer)
o Selectivity filter- evolutionary conserved structureo K+ channel selectivity filter catalyzes dehydration of ions, which confers specificity and speeds
up ion permeation
o How it works:
-
7/29/2019 Lipids, Membranes and Transport Outline
21/25
Lipids, Membranes and Transport Outline
21 | P a g e
P5 is where the K+ hydrated ion enters and in here, it dehydrates the K+ ion and allows it to fit specifically into
P4 to P0
Gating of K+ Channelso Gating means being able to shut if off (plug it up)o Different gating mechanisms define functional classes of K+ channelso Distinct from the selectivity filtero Regulated by the membrane potential
Packets
0,P1,P2,P3,P4,P5
Configurations
1,3 or
2,4
Keeps moving K+
up one rung-
every other, so it
pushes up, itsstaggered every
other one.
-Because positive
charges repel,
repulsion moves
cells up the
channel
Pushes up by repulsion
Opens when K+ is
resent
Pinch closed at bottom
Moves paddles, inverts
Able to close around
inside of channel
-
7/29/2019 Lipids, Membranes and Transport Outline
22/25
Lipids, Membranes and Transport Outline
22 | P a g e
Aquaporin channelso Selective water transport occurs through Aquaporin Channelso Have to dehydrate ions to move them througho Allow rapid and selective water transport across cell membraneso Tetramers of four identical subunits, with each subunit forming a pore
Aquaporin Selectivity Filtero Has 3 major features that confer a high degree of selectivity for water
Size restriction Electrostatic repulsion Water dipole orientation
o Water molecules have to line up single file based on dipole momentso Tetomer- forms like an hourglass, thin in the middle
Action Potentialso Electrical Signalso Depend on several typs of ion channelso Enable rapid communication between cells
Nerve cells and muscle cells use theseo Na+, K+, Ca2+ currents are key elements of action potentials, but not the only oneso One way that cells communicate with each other (rapidly)o Uses electrochemical gradientso Resting Potential- The electrical potential that exists due to a difference in charge across a
membrane. The charge thats maintained so the signal can be transmitted
Resting Potential- forces K+
against its gradient
+ charge outside, - inside
Voltage gated channels-
-Triggers gated ion channels to
open, triggers others to open up
as well
Action potentials are mediated
by ion currents- depends on ion
concentrations
-
7/29/2019 Lipids, Membranes and Transport Outline
23/25
Lipids, Membranes and Transport Outline
23 | P a g e
Action Potentialso Membrane depolarization is mediated by the flow of Na+ ions into cells through voltage
dependent Na+ channels
o Repolarization is shaped by transport of K+ ions through several different types of K+ channelso Electrical activity of organs can be measured as the sum of action potential vectors
Na+ Gradiento Used for getting lots of stuff into the cell
If we want to get something into the cell we can use Na+ ion to couple it, to get it inside to Transmembrane Na+ gradient is essential for the function of many transporterso Plasma membrane Na+ gradient is maintained by the action of the Na+/K+-ATPaseo Used for Glucose and other ionso Na+/K+-ATPase is an ion driven pump
ATP-driven Ion Pumps Generate Ion Electrochemical Gradientso Na+/K+-ATPase
Actively transport Na+ and K+ against their gradients by using the energy from ATPhydrolysis (to ADP)
3 Na+ exported for 2 K+ imported into the cell Antiporter- different directions Electrogenic pump- export 1 net positive charge (3 ions out, 2 ions in)
-
7/29/2019 Lipids, Membranes and Transport Outline
24/25
Lipids, Membranes and Transport Outline
24 | P a g e
Reaction cycle is described by the Post-albers scheme, which proposes that the enzymecycles between two fundamental conformations
Model for transport by t
Na+/K+-ATPase
1. 3 Na+ ions bind
a. ATP to ADP, and th
protein now has a
phosphate
2. Conformational chang
of molecule (protein)
3. Release 3 Na+ ions
outside the cell and its
opened up to allow K+
4. 2 K+ binds into pocke
protein gets
dephosphorylated
5. Protein closes outside
cell portion, protein gets
phosphorylated with AT
6. Releases K+ ions into
the cell
Starts all over again
-
7/29/2019 Lipids, Membranes and Transport Outline
25/25
Lipids, Membranes and Transport Outline
25 | P
Na+ gradient formed by Na+/K+-ATPase powers the transport of:o Examples
Na+/H+ and Na+/HCO3-cotransporters Regulates cytostolic and extracellular pH
Na+/Glucose transporter Intestinal cells absorb glucose
Na+/K+/Cl- cotransporter Regulates intracellular Cl- concentrations
Na+/Mg2+ exchanger Transport Mg2+ outside of the cell
Na+/Ca2+ exchanger Major transport mechanism for removal of Ca2+ from the cytosol of excitable
cells