Lipids and Membranes

Lipids Lipids are compounds that are soluble in non-polar organic solvents, but insoluble in water. Can be hydrophobic or amphipathic

Major Lipid Classes Acyl-lipids - contain fatty acid groups as main non-polar group Isoprenoids – made up of 5 carbon isoprene units

Lipid Subclasses

Function of major acyl-lipids Phospholipids – membrane components Triacylglycerols – storage fats and oils Waxes – moisture barrier Eicosanoids – signaling molecules (prostaglandin) Sphingomyelins – membrane component (impt. in mylein sheaths) Glycospingolipids – cell recognition (ABO blood group antigen)

Function of major isoprenoid lipids Steroids (sterols) – membrane component, hormones Lipid Vitamins – Vitamin A, E, K Carotenoids - photosynthetic accessory pigments Chlorophyll – major light harvesting pigment Plastoquinone/ubiquinone – lipid soluble electron carriers Essential oils – menthol

Fatty acids Amphipathic molecule Polar carboxyl group Non-polar hydrocarbon tail Diverse structures (>100 different types) Differ in chain length Differ in degree of unsaturation Differ in the position of double bonds Can contain oxygenated groups

Fatty acid nomenclature Short hand nomenclature describes total number of carbons, number of double bonds and the position of the double bond(s) in the hydrocarbon tail. C18:1 D9 = oleic acid, 18 carbon fatty acid with a double bond positioned at the ninth carbon counting from and including the carboxyl carbon (between carbons 9 and 10)

Fatty acid nomenclature Omega (w) notation – counts carbons from end of hydrocarbon chain. Omega 3 fatty acids advertised as health promoting Linoleate = 18:3 D9,12,15 and 18:3w3,6,9

Common saturated fatty acids common name IUPAC name melting point (Co) 12:0 laurate dodeconoate 44 14:0 myristate tetradeconoate 52 16:0 palmitate hexadeconoate 63 18:0 stearate octadeconoate 70 20:0 arachidate eicosanoate 75 22:0 behenate docosanoate 81 24:0 lignocerate tetracosanate 84

Common unsaturated fatty acids common name IUPAC name melting point (Co) 16:0 palmitate hexadeconoate 63 16:1 D9 palmitoleate cis-D9-hexadeconoate -0.5 18:0 stearate octadeconoate 70 18:1 D9 oleate cis-D9- octadeconoate 13 18:2 D9,12 linoleate cis-D9,12- octadeconoate -9 18:3 D9,12,15 linolenate cis-D9,12,15- octadeconoate -17 20:0 arachidate eicosanoate 75 20:4 D5,8,11,14 arachindonate cis- D5,8,11,14-eicosatetraenoate -49

Physical Properties of Fatty acids 18:0 18:1 18:3 Saturated chains pack tightly and form more rigid, organized aggregates Unsaturated chains bend and pack in a less ordered way, with greater potential for motion 70o 13o -17o

Melting points of fatty acids affect properties of acyl-lipids Membrane fluidity determined by temperature and the degree of fatty acid unsaturation of phospholipids Certain bacteria can modulate fatty acid unsaturation in response to temperature Difference between fats and oils Cocoa butter – perfect melt in your mouth fat made of triacylglycerol with 18:0-18:1-18:0 fatty acids Margarine is hydrogenated vegetable oil. Increase saturation of fatty acids. Introduces trans double bonds (thought to be harmful)

Unusual fatty acids can function analogously to unsaturated fatty acids

Major acyl-lipids Phospholipids – membrane components Triacylglycerols – storage fats and oils Waxes – moisture barrier Eicosanoids – signaling molecules (prostaglandin) Sphingomyelins – membrane component (impt. in mylein sheaths) Glycospingolipids – cell recognition (ABO blood group antigen)

Phospholipids Phospholipids are built on glycerol back bone. Two fatty acid groups are attached through ester linkages to carbons one and two of glycerol. Unsaturated fatty acid often attached to carbon 2 A phosphate group is attached to carbon three A polar head group is attached to the phosphate (designated as X in figure)

Common membrane phospholipids

Enzymes used to Dissect Phospholipid Structure

Plasmalogens Plasmalogens have hydrocarbon at carbon 1 attached thru vinyl ether linkage Polar head group could be ethanolamine or choline Important component of membranes in central nrevous system

Sphingolipids Sphingolipids named from Sphinx due to mysterious role Abundant in eukaryotic membranes, but not found in bacteria Structural backbone made of sphingosine Unbranched 18 carbon alcohol with a trans double bond between C4 and C5 Contains an amino group attached to C2 and hydroxyl groups on C1 and C3

Ceramides Sphingosine with fatty acid attached to carbon 2 by amide linkage Metabolic precursors to sphingolipids

Sphingomyelin has phosphocholine group attached to C1 of ceramide. Resembles phosphatidylcholine Major component of myelin sheaths that surround nerve cells

Cerebrosides contains one monosaccharide residue attached to C-1 of ceramide Glucose and galtactose are common Can have up to 3 more monosaccharide residues attached to sugar on C1 Abundant in nerve tissue Up to 15% of myelin sheath made up of cerebrosides

Gangliosides Gangliosides have oliosaccharide containing N-acetylneuraminic acid attached to C1 of ceramide Diverse class of sphingolipid due to variety of olgosaccharide species attached Oligosaccharide moiety present on extracellular surface of membranes ABO blood group antigens are gangliosides Impt in cell recognition, cell-cell communication

Defects in sphingolipid metabolism lead to disease state Tay-Saachs disease is a genetic defect in gangliosides degradation. Gangliosides accumulate in spleen and brain. Leads to retardation in development, paralysis, blindness, and early death. Niemann-Pick disease is a genetic defect in sphingomyelin degradation. Causes Sphingomyelin accumulation in brain, spleen and liver. Causes mental retardation. Children die by age 3 or 4.

Triacylglycerols (TAG) Fats and oils Impt source of metabolic fuels Because more reduced than carbos, oxidation of TAG yields more energy (16 kJ/g carbo vs. 37 kJ/g TAG) Americans obtain between 20 and 30% of their calories from fats and oils. 70% of these calories come from vegetable oils Insulation – subcutaneous fat is an important thermo insulator for marine mammals

Olestra Olestra is sucrose with fatty acids esterified to –OH groups digestive enzymes cannot cleave fatty acid groups from sucrose backbone Problem with Olestra is that it leaches fat soluble vitamins from the body

isoprenoids Isoprenoids are derived from the condensation of 5 carbon isoprene units Can combine head to head or head to tail Form molecules of 2 to >20 isoprene units Form large array of different structures

Terprenes

Steroids Based on a core structure consisting of three 6-membered rings and one 5-membered ring, all fused together Triterpenes – 30 carbons Cholesterol is the most common steroid in animals and precursor for all other steroids in animals Steroid hormones serve many functions in animals - including salt balance, metabolic function and sexual function

cholesterol Cholesterol impt membrane component Only synthesized by animals Accumulates in lipid deposits on walls of blood vessels – plaques Plaque formation linked to cardiovascular disease

Steroids

Many steroids are derived from cholesterol

Membranes Barrier to toxic molecules Help accumulate nutrients Carry out energy transduction Facilitate cell motion Modulate signal transduction Mediate cell-cell interactions

The Fluid Mosaic Model The phospholipid bilayer is a fluid matrix The bilayer is a two-dimensional solvent Lipids and proteins can undergo rotational and lateral movement Two classes of proteins: peripheral proteins (extrinsic proteins) integral proteins (intrinsic proteins)

The Fluid Mosaic Model

Motion in the bilayer Lipid chains can bend, tilt and rotate Lipids and proteins can migrate ("diffuse") in the bilayer Frye and Edidin proved this (for proteins), using fluorescent-labelled antibodies Lipid diffusion has been demonstrated by NMR and EPR (electron paramagnetic resonance) and also by fluorescence measurements Diffusion of lipids between lipid monolayers is difficult.

fusion After 40 minutes

Flippases Lipids can be moved from one monolayer to the other by flippase proteins Some flippases operate passively and do not require an energy source Other flippases appear to operate actively and require the energy of hydrolysis of ATP Active flippases can generate membrane asymmetries

Membranes are Asymmetric In most cell membranes, the composition of the outer monolayer is quite different from that of the inner monolayer

Membrane Phase Transitions Below a certain transition temperature, membrane lipids are rigid and tightly packed Above the transition temperature, lipids are more flexible and mobile The transition temperature is characteristic of the lipids in the membrane

Phase Transitions Only pure lipid systems give sharp, well-defined transition temperatures Red = pure phospholipid Blue = phopholipid + cholesterol

Structure of Membrane Proteins Integral (intrinsic) proteins Peripheral (extrinsic) proteins Lipid-anchored proteins

Peripheral Proteins Peripheral proteins are not strongly bound to the membrane They can be dissociated with mild detergent treatment or with high salt concentrations

Integral Membrane Proteins Integral proteins are strongly imbedded in the bilayer They can only be removed from the membrane by denaturing the membrane (organic solvents, or strong detergents) Often transmembrane but not necessarily Glycophorin, bacteriorhodopsin are examples

Seven membrane-spanning alpha helices, connected by loops, form a bundle that spans the bilayer in bacteriorhodopsin. The light harvesting prosthetic group is shown in yellow. Bacteriorhodopsin has loops at both the inner and outer surface of the membrane. It displays a common membrane-protein motif in that it uses alpha helices to span the membrane.

Lipid-Anchored Proteins Four types have been found: Amide-linked myristoyl anchors Thioester-linked fatty acyl anchors Thioether-linked prenyl anchors Glycosyl phosphatidylinositol anchors

Amide-Linked Myristoyl Anchors Always myristic acid Always N-terminal Always a Gly residue that links

Thioester/ester-linked Acyl Anchors Broader specificity for lipids - myristate, palmitate, stearate, oleate all found Broader specificity for amino acid links - Cys, Ser, Thr all found

Thioether-linked Prenyl Anchors Prenylation refers to linking of "isoprene"-based groups Always Cys of CAAX (C=Cys, A=Aliphatic, X=any residue) Isoprene groups include farnesyl (15-carbon, three double bond) and geranylgeranyl (20-carbon, four double bond) groups

Glycosyl Phosphatidylinositol Anchors GPI anchors are more elaborate than others Always attached to a C-terminal residue Ethanolamine link to an oligosaccharide linked in turn to inositol of PI

Membrane transport Membranes are selectively permeable barriers Hydrophobic uncharged small molecules can freely diffuse across membranes. Membranes are impermeable to polar and charged molecules. Polar and charged molecules require transport proteins to cross membranes (translocators, permeases, carriers)

Transport of non-polar molecules Non-polar gases, lipids, drugs etc… Enter and leave cells through diffusion. Move from side with high concentration to side of lower concentration. Diffusion depends on concentration gradient. Diffusion down concentration gradient is spontaneous process (-DG).

Transport of polar or charged compounds Involves three different types of integral membrane proteins Channels and Pores Passive transporters Active transporters Transporters differ in kinetic and energy requirements

Channels and Pores Have central passage that allows molecules cross the membrane. Can cross in either direction by diffusing down concentration gradient. Solutes of appropriate size and charge can use same pore. Rate of diffusion is not saturable. No energy input required

Porins Present in bacteria plasma membrane and outer membrane of mitochondria Weakly selective, act as sieves Permanently open 30-50 kD in size exclusion limits 600-6000 Most arrange in membrane as trimers

Passive Transport (Facilitated Diffusion) Solutes only move in the thermodynamically favored direction But proteins may "facilitate" transport, increasing the rates of transport Two important distinguishing features: solute flows only in the favored direction transport displays saturation kinetics

Three types of transporters Uniporter – carries single molecule across membrane Symport – cotransports two different molecules in same direction across membrane Antiport – cotransports two different molecules in opposite directions across membrane.

Saturation Kinetics of transport Rate of diffusion is saturable. Ktr = [S] when rate of transport is ½ maximun rate. Similar to M-M kinetics The lower the Ktr the higher the affinity for substrate.

Transporters undergo conformational change upon substrate binding Allows substrate to transverse membrane Once substrate is released, transported returns to origninal conformation.

Active Transport Systems Some transport occur such that solutes flow against thermodynamic potential Energy input drives transport Energy source and transport machinery are "coupled" Like passive transport systems active transporters are saturable

Primary active transport Powered by direct source of energy(ATP, Light, concentration gradient) Secondary active transport Powered by ion concentration gradient. Transport of solute “A” is couple with the downhill transport of solute “B”. Solute “B” is concnetrated by primary active transport.

Na+-K+ ATPase Maintains intracellular Na low and K high Crucial for all organs, but especially for neural tissue and the brain ATP hydrolysis drives Na out and K in

Na+-K+ ATPase Na+ & K+ concentration gradients are maintained by Na+-K+ ATPase ATP driven antiportsystem. imports two K+ and exports three Na+ for every ATP hydrolyzed Each Na+-K+ ATPase can hydrolyze 100 ATPs per minute (~1/3 of total energy consumption of cell) Na+ & K+ concentration gradients used for 2o active transport of glucose in the intestines

1o active transport of Na+ glucose

Transduction of extracellular signals Cell Membranes have specific receptors that allow cell to respond to external chemical stimuli. Hormone – molecules that are active at a distance. Produced in one cell, active in another. Neurotransmitters – substances involved in the transmission of nerve impulse at synapses. Growth factors – proteins that regulate cell proliferation and differentiation.

External stimuli(first messenger) – (hormone, etc…) Membrane receptor – binds external stimuli Transducer – membrane protein that passes signal to effector enzyme Effector enzyme – generates an intracellular second messenger Second messenger – small diffusible molecule that carrier signal to ultimate destination

G-Proteins Signal transducers. Three subunits, (a,b, g) a and g anchored to membrane via fatty acid and prenyl group Catalyze hydrolysis of GTP to GDP. GDP bound form is inactive/GTP bound form active When hormone bound receptor complex interacts with G-protein, GDP leaves and GTP binds. Once GTP -> GDP G-protein inactive GTP hydrolysis occurs slowly (kcat= 3min-1) good timing mechanism

Epinephrine signaling pathway Epinephrine regulation of glycogen degradation Fight or Flight response Ephinephrine primary messenger G-protein mediated response. G-protein activates Adenyl-cyclase to produce cAMP cAMP is the second messenger Activates protein kinase Activates glycogen phosphorylase

Effect of Caffeine Caffeine inhibits cAMP phosphodiesterase , prevents breakdown of cAMP. Prolongs and intensifies Epinephrine effect.

Phosphatidylinositol (PI) Signaling Pathway G-protein mediated G-protein activates phospholipase C (PLC) PLC cleaves PI to form inositol-triphosphate (IP3) and diacylglycerol (DAG) both act as 2nd messengers IP3 stimulates Ca2+ releases from ER DAG stimulates Protein kinase C