Lipid membrane
Learning outcomes • Describe the composition of the plasma membrane • Functions of the plasma membrane with reference to specific molecules • Fatty acid composition and functions • Describe phospholipid structure • Phospholipid functions
Membrane
Thickness: 7.5-10 nm
Membrane Functions 1. Compartmentalization (Intracellular compartments) 2. Scaffold for biochemical activity 3. Selectively-permeable barrier 4. Transporting solutes 5. Responding to external signals 6. Intercellular interactions 7. Energy transduction
The membrane is a lipid bilayer
Lipid bilayer comprises Phospholipid head groups (hydrophilic) And Associated fatty acids (hydrophobic)
e
Saturated C16 or C18 FA
Phosphodiester linkage
Derived from polar alcohol • smallest = H (from H-OH) • least common in membranes • phosphatidic acid
Unsaturated C16 – C22 FA
Saturated fatty acids - No double bonds - Space-saving - More rigid membranes Alpha linolenic acid
COOH CH3
Arachidonic acid
DHA
EPA
Unsaturated fatty acids - More than 1 double bond - Each double bond confers ‘kink’ - Occupies more space - Promotes fluidity ofmembranes
ω-3 fatty acids
ω-6 fatty acids
α-linolenic (18:3)
linoleic (18:2) Δ-6-desaturase Δ-6-desaturase
octadecatetraenoic (18:4)
γ-linolenic (18:3)
elongase eicosatetraenoic (20:4)
dihomo-γ-linolenic (20:3)
Δ-5-desaturase eicosapentanoic (20:5)
arachidonic (20:4)
PG3 and LT5
PG2 and LT4
The Importance of the fatty acids • Determine the fluidity of the membrane length of chain (usually 18-22) Level of unsaturation (# of C=C bonds)
• Lipids rarely move from one layer to another • Lipids exchange places with their neighbors • Lipids rotate around their axis
Membranes need to be fluid to: Enable the membrane proteins to diffuse rapidly Allow distribution of lipids and proteins properly in the lipid bilayer Ensure membranes can fuse with one another when necessary
Cholesterol in the Membrane
• Cholesterol is present where unsaturated lipids predominate • It ‘fills the gaps’ and stabilizes the bilayer • It stiffens the bilayer ∴ decreases fluidity & permeability
Extracellular face
75%
Outer layer stiffened by cholesterol Inner layer (fluidity determined by PUFA)
5%
20%
Rigid membrane
Fluidity determined by fatty acid composition of phospholipid Saturated: Palmitic (C16:0), Stearic (C18:1) Common fatty acids
Fluid membrane
Unsaturated: Arachidonic, Docosahexaenoic AA (20:4) DHA (22:6)
Membranes are Asymmetrical Glycolipids are found only on the extracellular surface (Sugar added in the Golgi)
• Inner & outer surfaces have different lipids • Proteins in the bilayer have a specific orientation due to its function
Phospholipids Phospholipids are Amphipathic (Amphipathic molecules are mostly lipid-like in structure, but have a hydrophilic region at one end
Choline Ethanolamine Serine Inositol
BASE
Phosphatidylcholine Phosphatidylethanolamine Phosphatidylserine Phosphatidylinositol
Phosphatidylinositol bisphosphate (PIP2)
Glycerol backbone
Inositol trisphosphate
Phospholipases PLA1 PLA2 PLC PLD
PLA1 CH2O..Stearic Acid
CH2O..Stearic Acid
CHO..AA
CHOH
CH2O..P..Base
CH2O..P..Base
Phospholipid
SA
Lysophospholipid
CH2O..Stearic Acid
CH2O..Stearic Acid
CHO..AA
CHOH
PLA2
CH2O..P..Base
Phospholipid
AA
CH2O..P..Base
Lysophospholipid
CH2O..Stearic Acid
CH2O..Stearic Acid
CHO..AA
CHO..AA
CH2O..P..Base PIP2
CH2OH
PLC IP3
Diacylglycerol
Action of phospholipase D
Phosphatidylinositol bisphosphate (PIP2) Membrane
3 phosphate groups ie PIP2
PLC hydrolyses PIP2
2 second messengers are produced
PIP2 hydrolysis
- stimulated by ligand-receptor Interactions - G protein-mediated - Leads to increased intracellular calcium concentration
PKC isozymes and some roles of PKC 1. Conventional (require DAG, Ca2+ and Phospholipid) PKCα, PKCβ (I and II), PKCγ
2. Novel (require DAG but not Ca2+) PKCδ, PKCε (I and II), PKCη and PKCθ
3. Atypical (require neither DAG nor Ca2+) PKCι, PKCζ (I and II) and PK-N1, N2
PKC is a serine-threonine kinase Roles (tissue- and cell-specific) include Receptor desensitization Modulation of membrane structure events Regulation of transcription Modulation of immune responses Regulation of cell growth Learning and memory.
Some lipid molecules act as signalling molecules Diacylglycerol AA Ceramide
Production of arachidonic acid
ACTIONS OF AA Activates ion channels directly Activates PKC Precursor of many signalling lipids
Phospholipases A2 PLA2 – categorized in terms of localization and association with calcium. (20 types identified; 9 in humans)
Secretory
Cytosolic Ca2+-independent (cytoplasmic) PAF-acetyl hydrolyses Roles: Inflammation, cell death, cell signaling, maintenance of membrane phospholipids.
Secretory PLA2 (sPLA2):
Originally from snake and bee venom Found extracellular especially in damaged tissue 10 groups (at least) Histidine residue at active site Require Ca2+ for activity characterized by a serine residue Roles: Linked with rheumatoid arthritis*, atherosclerosis, CNS inflammation, inflammatory bowel disease, skin inflammation, cancer, asthma Causes lysis of gram-positive bacteria *sPLA2-IIA primarily implicated in RA Released by macrophages Functions as a bacteriocidal
Cytosolic PLA2 (cPLA2):
4 groups (IVA, IVB, IVC and IVD named α, β, γ, δ). Active site is characterized by a serine residue. Ca2+-dependent Method of action: Ca2+-bound phosphorylated PLA2 translocates to intracellular membrane, cleaves phospholipid and releases arachidonic acid Pathological action (of head group/AA) can result in atherosclerosis, neuronal damage, multiple sclerosis, alziehmers, epilepsy. Groups IVA-C - high levels associated with colorectal, small bowel, and lung cancers
Ca2+-independent PLA2 (iPLA2):
Group members: VIA-1, VIA-2, VIB Active site characterized by a serine residue MW: 63 - 90 kDa (comparable in size to cPLA2). Cytoplasmic Roles: Maintenance of homeostasis through remodeling of membrane phospholipids Involvement in apoptosis, muscle contraction, and obesity. No direct links to inflammation and role in cancer not clear
PLA2 and inflammation
Arachidonic acid and metabolites
NOTES Arachidonic acid (C20:4) – 4 double bonds and therefore highly reactive Activates PKC Affects ions channels particularly K+ channels Affects transmitter release Double bonds mean it is a target for oxygenases Cyclooxygenases Lipoxygenases Epoxygenase
1) Cyclooxygenase 1 and 2: • Converts arachadonic acid into cycloperoxides • Cycloperoxides can then form thromboxanes, prostaglandins, and prostacyclins. • Ultimately roles of molecules include cell death, inflammation, vasoconstriction, and vasodilation in platelets, the endothelium, and in smooth muscle. • COX inhibitors eg VIOXX may be associated with increased risk of stroke. • Abnormal COX activity linked with cancer (2) Lipoxygenase (LOX): Converts arachidonic acid into hydroperoxyeicosatetraenoic acid (HPETE) and subsequently leukotrienes. (Leukotrienes have roles in vascular function and inflammation) Abnormal LOX expression has been linked with cancers, including colon, pancreatic, lung, prostate, bladder, skin, and liver cancer (3) Cytochrome P450 monooxygenase (CYP450): Converts arachidonic acid into 2 products, hydroxyeicosatetraenoic acid (HETE) and epoxyeicosatrienoic acid (EET). These molecules are peroxisomal proliferation activated receptor agonists (PPAR) Roles in angiogenic and mitogenic signaling. PPAR PPAR receptor is located at the nuclear membrane and dimerizes with 9-cis retiniose acid receptor following ligand binding; this causes binding to DNA at PPAR response elements. PPAR response elements are located near genes involved in lipid metabolism 3 types PPAR receptors: alpha, beta, and gamma. PPAR gamma is a modulator of the inflammatory response in peripheral macrophages, and monocytes (8).
AA gives rise to prostaglandins 1. NA, thrombin and bradykinin activate PLA2 leading to AA 2.
AA metabolites include prostaglandins thromboxanes prostacyclin
3. PGH2 is parent prostaglandin 4. Subsequent metabolism is enzymedriven
Some enzymes
PGE2 is synthesized from PGH2 by PGE synthase (Several types; PGES1 is key enzyme). PGD2 is synthesized from PGH2 by PGD2 synthases (2 forms known). PGI2 is synthesized from PGH2 via prostacyclin synthase PGF2a is synthesized from PGH2 by PGF synthase
PGs interact with specific receptors (pre- or postsynaptic) in brain PGD2 interacts with DP1, 2 PGE2 interacts with EP1-4 PGF2 interacts with FPα, β
PGI2 interacts with IP Thromboxanes interact with TPα, β
PG Receptors
Modulate physiological and pathological cellular functions
AA gives rise to leukotrienes
NOTES AA is omega-6 fatty acid DHA is omega-3 fatty acid
Omega-3 fatty acids
α-Linolenic acid (ALA) is parent fatty acid EPA and DHA are precursors for lipid modulators ALA .. plant sources (flaxseed, walnuts, pecans, hazelnuts, and kiwifruit) EPA and DHA … salmon, tuna, and herring
Omega-3 fatty acids
Linoleic acid (LA) is parent fatty acid Most from vegetable oils (soybean oil, corn oil, borage oil, and acai berry) LA converted to γ-linolenic acid (GLA) GLA converted to arachidonic acid
Lipid Analysis
Technologies: Analytical techniques for fatty acids, phospholipids, sphingolipids, triglycerides and steroids. Originally Thin layer chromatography (TLC) was used. Now replaced by faster technologies with better resolution. Gas chromatography-Mass Spectrometry (GC-MS) Used for low molecular weight lipids of all different classes (