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![Lipoprotein (a)
• Lp(a) has very similar structure as LDL
• it contains additional apoprotein molecule (a) [apo(a)], that is
covalently attached to apo B-100
• apo(a) has homologous structure with plazminogen
• large quantities Lp(a) in plasma are associated with the increased
risk of coronary heart disease
• hypothesis: Lp(a) slows down the breakdown of blood clots that
trigger heart attack because it competes with plasminogen for
binding to fibrin
67](https://image.slidesharecdn.com/1-lipoproteins-110920060058-phpapp01/85/1-lipoproteins-67-320.jpg)
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1 lipoproteins
- 1.
- 2. Recommended literature • R.K.Murray, D.K.Granner, V.W.Rodwell Harper‘s Illustrated Chemistry 27th, 28th Ed. • Harvey R.A., Ferrier D.R : Biochemistry 4th ed. • M.Lieberman, A.Marks: Basic medical Biochemistry (Clinical approach) 3rd edition • Silbernagl, Stefan; Despopoulos, Agamemnon: Color atlas of physiology, 6th ed. Published Stuttgart: Thieme 2009. • Lectures: – materials on IS MUNI 2
- 3. Lipids. Digestion and absorption,blood plasma lipids, lipoproteins 2011 Department of Biochemistry FM MU (E.T.) 3
- 4. Digestion of lipids Western diet contains 40 % of lipids or more. Triacylglycerols (TG)-90% Phospholipids (PL) Cholesterolesters (CE) Lingual and Glycolipids (GL) gastric lipase Lipophilic vitamins (LV) pancreatic lipases Bile acids + colipase Primary products: Absorption into mucosal free FA 2-monoacylglycerols cells (enterocytes) in lysophospholipids form of micelles (particles cholesterol < 20 nm) lipophilic vitamins 4
- 5. Cleavage of lipidsby enzymes in the small intestine • Pancreatic lipase Triacylglycerol 2-monoacylglycerol + 2 FA O O CH2 O C C O CH CH2 O C O < 1/4 TG triacylglycerol glycerol + FA 5 Orlistat - drug designed to treat obesity, it inhibits lipases
- 6. Orlistat 6
- 7. • Phospholipase A2 phospholipid lysophospholipid + MK O O CH2 O C C O CH O CH2 O P O CH2 CH2 NH2 O • Cholesterol esterase: Cholesterol esters cholesterol + FA 7
- 8. Emulsification of lipids •is condition for effective digestion of hydrophobic lipids • increase of effective surface oil-water, facilitation of contact with enzymes • is accomplished by action of detergents and mechanical mixing due to peristalsis Emulsificators in the small intestine • salts of bile acid • phospholipids • salts of fatty acids 8
- 9. Colipase • protein secretedfrom pancreas • binds the lipase at ratio 1:1 • anchors lipase to bile acids on the surface of emulsified lipid droplets • facilitates the action of lipase bile acids triacylglycerols colipase lipase 9
- 10. Absorption of lipidsby mucosal cells (enterocytes) Brush border Mucosal cells Mixed micelles microvilli diameter < 20 nm 10 Pasive difusion of fatty acid and monoacylglycerols
- 11. Absorption of lipidsby mucosal cells– cont. • long fatty acids and monoacylglycerol are resorbed by diffusion •short chain fatty acids (up to 10 C) do not enter micelles • they are resorbed directly • bile acids which remain in the intestine, are extensively absorbed in ileum • transport of cholesterol is mediated by NPC1L1 (Nieman- Pick C1 like 1) (see the lecture in Biochemistry I) 11
- 12. Hormons affecting digestionof lipids Secretin • intestinal “S-cells“ produce secretin into the blood after the stimulation by H+ entering the lumen • secretin stimulates release of secrets containing HCO3- from gallbladder and pancreas Cholecystokinin (CCK) • intestinal “I-cells“ produce cholecystokinin into the blood after the stimulation of small peptides and lipids • CCK stimulates secretion of amylase, lipase a proteases from exocrine cells of pancreas • potentiates effect of secretin on excretion of HCO3- • stimulates the secretion of bile from gallbladder 12
- 13. Steatorrhoea (lipid malabsorption) Loss of lipids by feces (normaly is resorbed ~ 98% lipids of food) Posible causes lipids Unsufficient supply of bile (damage of liver, obstruction of bile duct Disturbed function of pancreas Disturbed function of intestinal mucose Lipid Unsufficient intake of lipophilic malabsorption vitamins 13
- 14. Resynthesis of lipidswithin the mucosal cells: 1. Activation of FA FA + CoA + ATP Acyl-CoA + AMP + PPi 2. Resynthesis of triacylglycerols Acyl-CoA + Monoacylglycerol Diacylglycerol + CoA Acyl-CoA + Diacylglycerol Triacylglycerol + CoA 3. Resynthesis of phospholipids from lysophospholipids 4. Resynthesis of cholesterolesters Processes are located in ER FA with short chain and free glycerol do not také part in these processes and 14 are transported directly into the portal vein
- 15. Transport of lipidsfrom enterocytes AA apoprotein B-48 (apoprotein A-I) CHE PL TG chylomicron FA with short chain glycerol Lymphatic Portal vein lacteals 15
- 16. Plasma lipids Transport in form of lipoproteins Lipid Plasma concentration (mmol/l) Triacylglycerol 0,9 - 2 Total cholesterol 3,8 - 5,8 Non-esterified cholesterol 1,3 - 1,9 Total phospholipids 1,8 - 5,8 Free fatty acids 0,4 - 0,8 16
- 17. Schematic structure oflipoproteins Apoproteins Phospholipids Triacylglycerols Apoproteins Cholesterol esters Non-esterified cholesterol core superficial layer 17
- 18. Plasma lipoproteins TG 84% PL 54% CH Prot Chylomicron CM VLDL 45% 50% LDL HDL 18
- 19. Lipoproteins characteristics Class Diameter Half-life Main lipid (nm) CM 100-1000 ~5-15 min TG VLDL 30-90 ~2h TG IDL 25-35 ~2h TG/CHE LDL 20-75 ~2-4 d CHE HDL 5-12 ~10 h PL/CHE Separation of lipoproteins • ultracentrifugation in gradient of salt • elektrophoresis (see seminars and practicals) 19
- 20. Apoproteins • protein componentof lipoproteins • function of AP: enzyme activators and inhibitors interaction with receptors structural role (transport) • some of them are built in the lipoproteins, the other are peripheral and exchange between particles • synthesis in rough ER, attachment to lipid micelles 20
- 21. Chylomicrons (CM) –asembly and metabolism • they are formed in intestinal mucosal cells • they carry TG, CH and lipophilic vitamins admitted in food • main apoprotein is apo-B 48, minor is apo-A (the other cannot be synthesized in intestinal cells), synthesis of apoB-48 limits formation of CM • they are relased by exocytosis into the lacteals (lymphatic vessels originating in the villi of the small intestine) – chyle lymph • they follow lymphatic veins and enter the blood in the thoracic duct 21
- 22. Assembly of chylomicrons AA apoprotein B-48 (apoprotein A-I) CHE PL TG chylomicron SCFA glycerol Lymphatic Portal vein vessels 22
- 23. Metabolism of chylomicronsin blood • they enter the blood 1-2 hours after the meal – nascent chylomicrons • modification of chylomicrons: in blood apo E and apo C-II are transfered from circulating HDL to chylomicrons • in capilaries of most peripheral tissues are CM degraded by lipoproteine lipase (LPL) 23
- 24. Chylomicron in blood(1st part) blood ApoCII Ductus thoracicus ApoE Adipose lymph tissue, muscles LPL Fatty acids Mucosal cells of the intestine glycerol 24
- 25. Lipoprotein lipase (LPL) •negatively charged enzyme on the surface of endotelial cells in capillaries (anchored by heparansulfate to the capillary walls) • predominantly in adipose tissue and skeletal and cardiac muscle • it is activated by apo-CII • LPL can be released by heparin • synthesis of isoenzyme in adipose tissue is stimulated by insulin • deficit of LPL results in triacylglycerolemia 25
- 26. Action of LPL •lipoprotein is attached to the enzyme bonded on endothelial cells • LPL catalyzes hydrolysis of TG contained in circulating lipoproteins: TG glycerol + 3 MK O O CH2 O C C O CH CH2 O C O 26
- 27. Action of LPLon chylomicrons CM • free fatty acids are Apo CII taken up by tissues, small part returns back into the plasma and is LPL LPL transported by serum albumin • LPL hydrolyses TG to fatty acids and CM-remnat glycerol Is rapidly removed • more than 90% of TG in CM is from circulation by degraded by LPL, the particle decrases in the liver játry size and increases density remnant (Apo-E receptor) 27
- 28. Chylomicron in blood(2nd part) ApoCII ApoE Adipose tissue, muscles LPL ApoCII Fatty acids liver Glycerol apoE apoE, apoB/E, remnant 28
- 29. • LPL degradesabout 90% TG in chylomicrons • chylomicrons in blood are removed during aprox. 30 min The fate of fatty acids released by the action of LPL: - oxidation in tissues (muscle, myocard) – yield of energy • deposition in form of TG in adipous tissue Removal of remnats from blood •They bind to the receptors in liver and are taken up • receptors recognize apo-E • cholesterol taken in food is transported into the liver in 29 this way
- 30. VLDL – asemblyand metabolism • they are formed in hepatocytes • they are composed of 60% of TG that are synthesized in the liver • contains cholesterol • content of apoproteins:apo-B 100, small amount of Apo-A and Apo-C • they are secreted into the blood as nascent particles 30
- 31. Apo-B 100 • apoproteinin LDL and VLDL •It is integral protein • very long chain (4 536 AA) • apo-B 48 and apo-B 100 have the same mRNA, but apo- B 48 has only 48% of chain leghth of apo-B 100) • synthesis of apo-B 100 is inhibited by insulin, VLDL are formed in post-resorption phase 31
- 32. What is theorigin of TG in liver? • fatty acids are synthesized in liver from acetyl-CoA • acetyl CoA originates mainly from metabolism of saccharides (after the meal) • free fatty acids can be also taken up from blood (during starvation) • TG are synthesized from fatty acids 32
- 33. Metabolism of VLDL- overview VLDL Liver apoB/E Apo-C,E HDL Tissues LPA Apo C FA G IDL HDL LDL CHE 33
- 34. Degradation of VLDLby lipoprotein lipase Apo CII free fatty acids enter tissues LPL IDL particle become smaller in size LDL Taken up by liver (apo-E receptor) 34
- 35. Metabolism of VLDLin steps •Nascent VLDL enter the blood • apo E and apo CII are transfered from HDL • triacylglycerols are degraded by lipoprotein lipase to fatty acid and glycerol (similarly like chylomicrons) •VLDL changes to IDL • IDL are either taken up by liver or are converted to LDL 35
- 36. Hepatic lipase • enzymon luminal wall of liver sinusoids • it acts similarly like LPL • it degrades TG in IDL, VLDL and HDL when they pass through the liver • it can be released by heparin 36
- 37. Heart muscle andadipose LPL KM of heart muscle LPL is aprox. 10x lower than adipose LPL Synthesis of adipose LPL is activated by insulin What follows from it ? 37
- 38. Metabolism of IDLand LDL • IDL and LDL may be enriched by cholesterol esters from HDL (role of cholesterolester transfer protein CETP) • IDL are taken up by liver Apo-B/E receptors • LDL are taken up by periferal tissues (1/3) and liver (2/3) by the process of receptor mediated endocytoses (Apo-B/E) • at physiological conditions 30-40% of new formed LDL is catabolized during 24 h 38
- 39. LDL receptors LDL receptor(apo B/E Non specific (scavenger) receptor) receptors (SR-A) Is regulated by intracelular membrane receptor with broad content of cholesterol specifity Located on the surface of macrophages and Kupfer-cells in liver Are not down regulated Uptake of modified and redundant LDL 39
- 40. Uptake of LDLby specific receptors (apo B/apoE) •LDL receptor is negativelly charged membrane glycoprotein lokalized on the surface of clathrin coated pits, • it recognizes apo E and apo B 100 40
- 41. • after binding,the complex LDL-receptor is internalized by endocytoses 41
- 42. •vesicles containing LDLloses clatrin coat and fuses with lysosomes forming endosomes •LDL disociates from its receptor, the receptor migrates to one side of endosome, separates a recycles back to the membrane membrány recyclation of a receptoru Lysosome 42 endolysosom
- 43. •pH in endolysosomefalls, lipoprotein is degraded, cholesterol, amino acids, fatty acids, phospholipids releases • cholesterol is esterified by ACAT (acylCoA-cholesterol- acyltransferase) and is stored in the cell ACAT CH CHE AK 43 MK PL
- 44. The level ofcholesterol in the cell is strictly regulated • down- regulation of cholesterol intake in form of LDL (increased level of cholesterol in the cell decreases the number of receptors on the surface – expression of LDL-receptor gene is decreased) • regulation of intracellular synthesis of cholesterol regulation of transcription of HMG-CoA synthase gene by SREBP Inhibition of HMG-CoA – reductase by cholesterol 44
- 45. Role of sterolregulatory element-binding protein (SREBP) in regulation of intake and synyhesis of cholesterol • precursor of SREBP is an integral protein of ER membrane • when sterol level in the cell is low, N-terminal peptide is released from precursor molecule and migrates to the nucleus, where it binds to sterol-regulatory element in promotor area of genes regulating cholesterol • SREBP regulates synthesis of LDL receptors and HMGCoA reductase See also: http://www.biocarta.com/pathfiles/m_s1pPathway.asp 45
- 46. Familiar hypercholesterolemia (type IIhyperlipidemia) Defeciency in production of LDL-receptors High level of LDL in blood Synthesis of cholesterol in the cell is not inhibited cell produces excess of cholesterol Increased risk of myocardial Characteristic tendon infarction xanthomas 46
- 47. Uptake of lipoproteinsby scavenger receptor SRA • SRA –receptors on surface of phagycytosing cells (macrophages in the cell wall, in lung alveoles and peritoneum, Kupfer cells) • receptor does not have down-regulation • it preferably take up modified LDL (oxidized, glycated) • it can take up also undamaged LDL, if the capacity of down- regulated receptor is exceeded 47
- 48. Formation of foamcells and plaque •Makrophages filled by lipids become foam cells • they canoothcumulate in subendotellial area of the cell wall • growth factors and cytokines stimulate the migration of smooth muscle cells and their proliferation • these processes can result in formation of atherosclerotic plaque 48
- 49. Effect of hormoneson LDL uptake in liver Insulin and trijodthyronin increase uptake of LDL by the liver, Glucocortikoids have opposite effect (mechanism is not known) Why non-controlled diabetes and hypothyroidism are risk factors for development of atherosklerosis and are very often connected with hypercholesterolemia? 49
- 50. HDL and theirrole in metabolism of lipids reverse transport of cholesterol (RTC) - HDL take up cholesterol from peripheral tissues and transports it to the liver HDL exist in several modifications They differ in size, shape, by content of lipids and apoproteins • they have different functions • main subfraction according to the density: HDL2, HDL3 • remodelation of HDL – changes of HDL in circulation as 50 a consequence of exchange of lipids
- 51. HDL – assemblyand metabolism • base of HDL structure are apo A I and apo A II, they contain also apoC and apo E • for transport of cholesterol from tissues are important small particles so-called „lipid free“ a „lipid poor“ apo A • thet are secreted by liver and enterocytes, can be also formed in circulation from larger HDL particles 51
- 52. SR-B1 receptor HDL-receptor withdual function in metabolism of HDL • It binds HDL in liver and steroidogenic cells through apo A-I and mediates transport of cholesterol inside the cells • it mediates the transport of cholesterol from the cells into the HDL in periferal tissues 52
- 53. How is cholesteroldeposited in the cells? •Cholesterol esters are present in form of lipid droplets in cytoplasma •Revers transport of cholesterol is started by their hydrolysis by cholesterylester hydrolase •Free cholesterol is transported to the cell membrane 53
- 54. How is cholesteroltaken up from tissues ? Two mechanism are expected difussion formation of new HDL 54
- 55. Formation of newHDL Lipid poor Periferal particles tissue játra SR-B1 ABCA1 is an ATP- ABCA1 binding cassette steroidogenní tkáně Tangier disease – extremly rare genetic disease – LCAT CETP functional ABCA1 transporter is missing. It results in intracellular spherical HDL accumulation of lipids Disk-shaped HDL LDL/VLDL * Gene expression for pro ABCA1 is regulated by the amount of cholesterol 55 inside the cell
- 56. Mechanism of cholesteroldifusion into HDL •HDL attaches to cell surface after the interaction with SR- Periferal B1 receptor tissue •ABCG4 (ATP-binding casette protein G4) transports SRB-I cholesterol into the HDL ABCG4 •Gradient of cholesterol concentration is mediated by LCAT on the HDL surface 56
- 57. For further conversionsof HDL are important: Lecitin cholesterol acyltransferase (LCAT) Cholesterol ester transfer protein (CETP) LCAT • transfers a fatty acid from lecitine (phosphatidylcholine) na cholesterol • plasmatic enzyme, it acts on the surface of HDL, activated by apo A-I 57
- 58. Function of LCAT CH2 O CO CO O CH O CH3 + CH2 O P O CH2 CH2 N CH3 O- CH3 • LCAT transfers acyl of fatty acid to OH group of cholesterol • non-esterified cholesterol is HO converted to esterified it is less polar and more voluminous – it is sequestered into the core of HDL 58
- 59. CETP Cholesterolester –transfer-protein It transfersesterified cholesterol from HDL to other lipoproteins CE VLDL CETP CE HDL 59
- 60. Further metabolism ofHDL • esterified cholesterol formed by the action of LCAT accumulates in the core of HDL • particles become spheric • spheric HDL interacts with the other lipoproteins • uptake in liver is mediated by SR-B I receptor 60
- 61. Cholesterol uptake bySR-B1 receptor in the liver •HDL binds to receptor on hepatocytes • the complex is not endocytosed • only cholesterol is transported into the cells, transport is mediated by ABCG transporter •HDL dissociates from receptor and re-enters again circulation 61
- 62. Conversions of HDL- summarization Lipid poor particles Peripheral játra SR-B1 tissue SR-B1 ABCA1 steroidogenic cells SRB-I LCAT CETP Disk-shaped spherical HDL LDL/VLDL (HDL2 a HDL3) HDL Remodelation by interaction with the other 62 lipid poor particles lipoproteins
- 63. Cholesterol balance Food Biosynthesis 80-500 mg 800 – 1000 mg Cholesterol pool Steroidal hormons, Cholesterol Bile acids sebum, intestinal (bile) epitelium 800 mg 500 mg 200 mg 1000-1500 mg/day is secreted 63
- 64. “Good” and “bad” cholesterol HO HO Cholesterol in HDL is considered to be the good cholesterol, because it accepts free cholesterol from peripheral tissue 64
- 65. Increased intake ofcholesterol or defects of LDL receptors Increased plasmatic LDL, The long half-life of LDL possibility of oxidation Damaged and redundant LDL are taken up by SRA receptors of makrophages, formation of foam cells oxidized LDL are strongly aterogenic LDL cholesterol – bad cholesterol 65
- 66. High level ofHDL-cholesterol • HDL act as cholesterol scavenger, picking up excess cholesterol in blood and taking it back to the liver for disposal. The higher HDL cholesterol level, the better prognosis of coronary heart diseases risk. HDL-cholesterol = good cholesterol When the higher level of cholesterol in blood is found, the distribution between LDL and HDL fractions is determined – examination of HDL- and LDL- cholesterol is performed – see practicals 66
- 67. Lipoprotein (a) • Lp(a)has very similar structure as LDL • it contains additional apoprotein molecule (a) [apo(a)], that is covalently attached to apo B-100 • apo(a) has homologous structure with plazminogen • large quantities Lp(a) in plasma are associated with the increased risk of coronary heart disease • hypothesis: Lp(a) slows down the breakdown of blood clots that trigger heart attack because it competes with plasminogen for binding to fibrin 67