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57 Cards in this Set
- Front
- Back
Separate cell contents from the extracellular environment
Separate organnels from other cellular constituents |
Membranes are biological barriers
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Membranes provide ________ and _____ the flow of metabolites and other compounds in/out of the cell.
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protection and control the flow of metabolites.
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Amphiphatic lipids, such as phospholipids and sphingolipids
These lipids contain both polar and non-polar components. When put in an aqueous medium, tend to self-associate into a spherical or a laminar structure. |
Membranes are based on lipids
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Has a hydrophobic core
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Micelle
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Lipid bilayer enclosing an aqueous core
Inject DNA into a cell |
Liposomes
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Membranes also contain proteins with a variety of functions
Carbohydrates can be bound to both the lipid and protein components Different membranes have different compositions of protein, lipid and carbohydrate Biological membranes are asymmetric |
Membrane composition
Cannot flip from inner to outer membrane |
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Carbohydrates are mostly located on the ______ surface of a cell membrane
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outer
Often serve as recognition sites for specific binding |
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Different compositions affect the membranes properties
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Lipid/protein ratio matters
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The inner and outer faces have different protein and lipid profiles.
Maintains entry into cells |
The asymmetry of phosphatidylserine contributes to the membrane potential due to its negative charged.
Phosphatidylserine is a marker for apoptosis An attempt to break voltage dependent linkages |
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Different membranes have different lipid and ______lipid compositions
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phospholipid
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Components can move laterally within a membrane face
Components move from one face to the other slowly, if at all. |
Asymmetry impleis that both lipid and protein are 'fixed' on a given face
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Fluidity is dependent upon ________ and _______
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temperature and composition
Membranes are fluid structures |
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Membranes have a transition temperature at which they undergo a phase change
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Fluidity increases with increasing temperature
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- Increased saturation of fatty acid components decreases fluidity
- Increased cholesterol decreases fluidity - Ca2+ binding to the ionic portion of lipid components can decrease fluidity. (provides chelator) |
Fluidity is affected by composition, particularly the lipid composition
Fatty acids like to congregate tightly. Unsaturated bonds form kinks and push them farther apart. |
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Polar head group associates with hydrophilic outer layer
Hydrophobic ring structure nestles into hydrophobic acyl chains. Heaad group can hydrogen bond with phospholipid head groups in the hydrophilic region |
Cholesterol
- Decreases fluidity caused by cis-double bonds - Gets between saturated bonds and prevents them from congregating Overall, fluidity decreases |
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- May contain hydrophobic transmembrane domains that traverse the membrane
- May be mostly on the inside or outside face of the membrane - May be covalently bound to a lipid - Removal requires drastic treatment (detergent, organic solvent), often with a loss of native structure |
Integral proteins are bound within the membrane, including the lipid portion
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- Mostly through polar interactions
- May be through interaction with an integral protein - Easily released by ionic solvents |
Peripheral proteins are bound to the surface of the membrane
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- The glycosylphosphatidylinositol (GPI) anchor
- Attachment of myristic acid (C14:0) by N-terminus - Attachment to FA (C14-18:0) by thioester bond with cysteine - Attachment to prenyl groups through thioester bond with cysteine |
Colvalent attachment of proteins to membrane lipids
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Conjugation of a protein to an inositol.
Essentially integrated into membrane. |
The glycosylphosphatidylinositol anchor (GPI anchor)
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1. Specific enzyme catalysts
a. ETC, succinate dehydrogenase, carnithine acyltransferase 2. Transport of solutes across membranes a. Transporters, translocases, translocators, permeases, pumps 3. Sites of recognition a. Usually through bound carbohydrate b. Surface immunoglobulins in cells of the immune system c. Peptide hormone receptors 4. Structural component a. Maintenance of cell shape or flexibility. b. Cytoskeleton c. Anchored to trans-membrane proteins |
Functions of proteins in membrane
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- Fluidity
- Flexibility - Self-sealing ability - Impermeability - Sidedness |
Characteristics of the Fluid Mosaic model membrane
Self-sealing: Molecules are separated, then heal/repair itself |
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Integral and peripheral proteins and cholesterol
Carbohydrate binding |
More properites of the FMM membrane
Implies sidedness. |
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Determined empiracally by measuring rates of lateral movement of specific lipids and other membrane components.
Observed their distributions |
The Lipid Raft model
Counters the FMM of lateral diffusion |
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Membranes contain heterogeneous dirstributions of lipids and proteins.
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Lipid Raft model
As opposed to homogenous distribution of the FMM |
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Domains with high concentrations of cholesterol, sphingolipids and localized concentrations of certain proteins.
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These domains are called: Lipid Rafts
Rafts are small: - Tens of microns in diameter - Don't comprise the majority of the membrane |
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Certain _______ proteins (eg, receptors) have been shown to localize lipid rafts.
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membrane proteins
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This ________ fence model proposes that there are isolation of membrane subdomains encircled by proteins.
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Picket fence
May involve the anchoring of these proteins to the cytoskeleton May result inphase transitions between domains, due to differing lipid and cholesterol composition. |
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Raft organization appears to be dynaming and perhaps regulated.
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Key point of Lipid Raft model
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Testing has shown that certain molecules prefer to target these areas of a cell
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Pockets of high cholesterol in a raft
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1. Electrochemical transport
a. Chemical i. Relative number of solutes on either side of a membrane b. Electrical ii. Inside of cell is negative, due to anions within the cell. 2. Passive transport 3. Active transport |
Mechanisms of transport across membranes
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Uncommon mechanism
Primarily for gases (CO2, O2) Not for charged molecules |
Simple diffusion (passive transport)
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- Specific
- Saturable - Inhibitable |
Mediated transport
Much more like an enzyme catalyzed reaction |
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- Uniport
- Symport Drags molecule against its concentration gradient, utilizes the high gradient of another molecule. Ex: glucose transport into intestinal epithelial cells. - Antiport Na+/K+ ATPase |
Types of mediated transport
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Energetics of transport
dG = 2.3 RT log [S2]/[S1] |
If dG is negative, transport is passive
If dG is positive, transport is active |
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Undergo a conformation change to move substrate across membrane
Functions like an enzyme |
Transporters
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Maintain an open cavity for passage of a substrate
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Channels/pores
Channel: Selective, usually gated. Behave more like transporters Pore: Non-selective, larger opening |
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Commonly formed by amphipathic alpha helices
Allow passage by opening/clsoing ('gating') based on conformation changes Gating may be dependent on: a. Membrane potential (voltage-gated channels) b. Binding of a specific agonist (ligand-gated channels) c. Covalent modification (eg phosphorylation) |
Ion channels
Clinical significance: Cystic fibrosis is a deficiency in a chloride channel. |
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A member of the ATP binding cassette (ABC) family of proteins
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CFTR found in epithelial cells of multiple organs
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Channel is only active when phosphorylated.
a. Phosphorylation of the regulatory subunit (R) by PKA causes a confirmation change that allows ATP to bind to the adenine nucleotide binding domain (ABD) to initiate channel opening. |
CFTR is a gated channel controlled by phosphorylation
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Mutations that affect CFTR function result in the accumulation of thickened mucus in airways, leading to pathologies associated with cystic fibrosis
Cholera toxin activates CFTR phosphorylation, causing constituitve channel opening. i. In the intestine, this causes dehydration. |
Chloride efflux is important for the osmotic movement of water out of the cell to maintain mucous fluidity
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CFTR required for osmotic balance in lung, liver, pancreas, GI tract, reproductive tract and skin.
CFTR defects can affect the function of any of these organ systems. CFTR deficiencies have been attributed to >1400 mutations. Most mutations result in unstable proteins that are degraded in the ER The most common mutation (66% of cases) is a deletion of Phe 508, which causes improper folding of CFTR and its degradation by the cell. Defects in CFTR cause varying degress of dysfunction of most epithelia. |
CFTR affects epithelium in multiple tissues.
Phe 508 - Not a dysfunctional protein - Protein isn't there |
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1. PKA phosphorylation of the RD renders NBDs available for ATP binding.
a. CFTR is unique among ABC transporters in containing an R-domain that controls ATP binding. 2. ATP binding causes NBD1/NB2 dimerization, and the conformational change that opens the channel. 3. Hydrolysis of ATP at site 2 releases of ADP, disrupting NBD dimerization and closing the channel. 4. The channel cycles approximately once per second. 5. Unclear how many Cl- ions pass through per cycle. |
CFTR mechanism in action
Coordinated ATP binding of both cells Hydrolysis of ATP and release of ADP: Disrupts NBD dimerization and closes the channel |
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ATP-binding cassette transporters: A superfamily of transporters in all organisms that move diverse compounds across membranes.
a. Driven by a cycle of conformational changes mediated by ATP binding (opens) and hydrolysis (closes). Most are active transporters, not gated channels like CFTR. Originally described as ATP-dependent nutrient transporters in bacteria. e.g. Maltose transporter Also function in bacteria to pump endogenous virulence factors and antimicrobial agents out of the cytoplasm. |
ATP-binding cassette transporters (ABC transporters)
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- In humans, ABC family comprised of 49 transporters in 7 subfamilies (based on structural similarities, designated ABCA through ABCG) with diverse functions in various cell types and organelles.
a. Regulate local permeability at blood barrier in brain, spinal cord, and placenta. b. Excrete toxins in the liver, GI and kidney. c. Important for cellular lipid transport and metabolism. d. Antigenic peptide processing/surface presentation in MHC. e. Can modulate lipid composition of outer and inner layers of the plasma membrane. f. Examples of ABC-transported molecules: cholesterol, lung surfactant, peptides,heme, bile salts, phospholipids, xenobiotics, toxins. |
ABC transporters
Of note, Antigentic peptide processing/surface presentation in MHC |
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Broad substrate specificity of prokaryotic transporters contributes to the acquisition of multiple drug resistance (MDR) by pathogenic bacteria.
Serious problem with nosocomial infections (e.g. MRSA). |
ABC transporters as drug pumps:
Multiple drug resistance (MDR) in bacteria & cancer cells |
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13 human ABC transporters have been associated with drug transport and resistance, primarily from the ABCC family.
Also designated as MRP (multidrug resistance protein) in this context. Overexpression causes broad specificity resistance to drugs, due to their rapid efflux from the cell. Can dramatically affect drug efficacy. Particulary problematic for cancer treatment by chemotherapeutics. Cancers can acquire this phenotype over type, due to the selection of drug resistant cells. Each can generally transport multiple unrelated drugs. Collectively shown to transport hundreds of different drugs compounds. |
ABC transporters as drug pumps:
Multiple drug resistance (MDR) in bacteria & cancer cells |
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First/best characterized example is ABC-B1 (P-gp/MDR1), associated with the acquisition of Multiple Drug Resistance (MDR) in many cancers.
Overexpression confers high resistance to many classes of drugs Anthracyclines, vinca alkaloids, taxanes, etoposide, imatinib, colchicine, methotrexate, actinimycin D, et al. |
ABC transporters as drug pumps:
Multiple drug resistance (MDR) in bacteria & cancer cells Research is now directed toward inhibition of these pumps. |
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1) Substrate binding induces a conformational change that enhances ATP binding to NBD site 1.
2) Binding of ATP to site 1 stabilizes NBD interaction, facilitating ATP binding to NBD site 2. 3) Binding of second ATP completes formation of the NBD dimer, and causes a conformational change in the MSDs that releases the substrate on the opposite side. 4) The protein maintains outward facing orientation as site 2 ATP is hydrolyzed. 5) Release of ADP from site 2 resets the protein for inside face substrate binding. Step 6? Not clear whether site 1 ATP hydrolysis and ADP release is also required to reset. |
Current model of the ABC/ MRP transport cycle
Example: MRP1 (ABCC1) and leukotriene C4 (LTC4) transport |
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The Na+/K+-ATPase contributes to the electronegativity of the cytosol
(i.e. the membrane potential) due to the net efflux of positive charges. |
Glucose transport through the cell
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Glucose exits to plasma by passive mediated transport by this transporter
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GLUT 2
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- K+ is transported out of the cell to make room for Na+ influx
- Helps maintain electronegativity |
Leakdown transporters (I.E. K+ transporters)
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- Intracellular compartmentation is a defining characteristic of eukaryotic organisms.
- Compartmentation allows for the restriction of processes and reactions to physically separate environments within the cell. a. Mediated by phospholipid membrane barriers between organelles and the cytoplasm. b. Permits optimal conditions for subsets of processes without conflict. i. Allows for disparate concentrations of molecules between compartments. c. Provides an additional mechanism for regulation by controlling the interaction between compartments. |
Compartmentation and shuttle properties
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The inner mitochondrial membrane contains numerous specific transmembrane shuttles for the selective movement of small molecules between the mitochondrial matrix and the cytosol.
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This is particularly important for intermediary metabolism with respect to the interaction between reactions that occur in the cytosol and those that occur in the motochondria.
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Citrate Transporter
Citrate generated in mitochondria is exported to cytosol where it serves as a source of acetyl-CoA, in exchange for malate. |
Metabolic pathways use multiple transporters
in shuttle systems |
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ADP/ATP and Phosphate Translocation
a. Adenine nucleotide transporter b. Phosphate translocator |
Metabolic pathways use multiple transporters
in shuttle systems |
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Glycerol Phosphate Shuttle & Malate/Aspartate Shuttle
Transport shuttles for reducing equivalents |
Metabolic pathways use multiple transporters
in shuttle systems |
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Acylcarnitine transport (CPT I & CPT II)
A transport shuttle for acyl-CoA for oxidation in mitochondria |
Metabolic pathways use multiple transporters
in shuttle systems |