Dr. M. Azhar Chishti Dept. Medical Biochemistry Lecture Objective 1. To study the structure of physiologically important phospholipid classes- glycerophospholipids and sphingolipids. 2. To discuss the synthesis of phospholipids of clinical and physiological relevance and their degradation by phospholipases. 3. To study the role of phospholipids like phosphatidylinositol in signal transduction and membrane anchoring choline, ethanolamine and serine, cardiolipin and sphingomyelin etc. Lecture Outline 1. Overview of phospholipids. 2. Structure of Glycerophospholipids and sphingolipids. 3. Synthesis of phospholipids: Phosphatidic acid, phosphatidyl-choline, ethanolamine and serine, cardiolipin and sphingomyelin etc. 4. Degradation of phospholipids by phosphalipases. 5. Role of phosphatidyl choline in lung surfactant 6. Role of phsophatidylinositol in signal transduction and membrane anchoring . Phospholipid Polar compounds composed of alcohol attached by a مربوط بـ رابطهphosphodiester bridge to either diacylglycerol or sphingosine. Amphipathic ) متردد ( محب وكاره للماءin nature, has a hydrophilic head (phosphate + alcohol eg., serine, ethanolamine, and choline) and a long, hydrophobic tail (fatty acids or derivatives ). In membranes, the hydrophobic portion is associated with the nonpolar portions such as glycolipids, proteins, and cholesterol. The hydrohilic polar head extends outward, facing intracellular or extracellular aqueous environment (see Figure). االسماء تشرح في شريحه 7 Membrane phospholipids also function as : 1- a reservoir حافظfor intracellular messengers, and, 2-for some proteins, phospholipids serve as anchors ) مرساه ( مثبتto cell membranes. Non-membrane-bound phospholipids الفوسفولبدز الذي لم يدخل في : تركيب الجدر الخلويه ( حر ) له وظيفهserve additional functions in the body, for example, 1-as components of lung surfactant and 2-essential components of bile, where their البايل هي المكان الذي له وظائف تنظيفdetergent properties aid "in the solubilization of cholesterol. مثل اخراج الكلويسترول ارتباطات الفسفولبدز بالجليسرول: اوال (PA) Glycerophospholipids : are formed from phosphatidic acid and an alcohol. Serine + PA -•phosphatidylserine (PS) Ethanolamine + PA -•phosphatidylethanolamine (cephalin) Choline + PA -•phosphatidylcholine (lecithin) PC Inositol + PA -•phosphatidylinositol (PI) Glycerol + PA•phosphatidylglycerol (PG) (Cardiolipin) اسم فسفولبدز يستخدم للدالله على مرض الزهري Two molecules of phosphatidic acid esterified through phosphate to an additional molecule of glycerol are called cardiolipin. This is the only human glycerophospholipid that is antigenic. cardiolipin is recognized by antibodies raised against Treponema the bacterium that causes syphylis الزهري. [Cardiolipin is an important component of the inner mitochondrial membrane and bacterial membranes.] Plasmalogens When the fatty acid at carbon 1 of a glycerolphospholipid is replaced by an unsaturated alkyl group attached by an ether ( رابطه ايثرrather than بدال منby an ester) linkage مهمهto the core glycerol : molecule, a plasmalogen is produced. Phosphatidalethanolamine is abundant غني بـin nerve tissue is a plasmalogen. Phosphatidalcholine (abundant in heart muscle) is the other quantitatively كميهsignificant ether lipid in mammals. Platelet-activating factor (PAF) PAF is an unusual ether مرتبط برابطه ايثر ايضاglycerophospholipid, with a saturated alkyl group ان لديها رابطه مشبعه-1تختلف عن اللي قبلin an ether link to carbon 1 and >>> االختالف الثاني2- an acetyl residue (rather than a fatty acid) at carbon 2 of the glycerol backbone. PAF is synthesized and released by a variety of cell types that binds to surface receptors, triggering potent قويthrombotic and acute inflammatory leads to hypersensitivity & anaphylactic ردود فعل مناعيهreactions. It causes platelets to aggregate and degranulate the neutrophils and alveolar macrophages to generate superoxide radicals that killed the bacteria. PAF is one of the most potent bioactive molecules known, causing effects at concentrations as low as 10 -12 mol/L. ارتباط الفسفولبدز بالسفينقو: ثانيا Sphingophospholipids or sphingomyelin The backbone of sphingomyelin is the amino alcohol sphingosine, rather than glycerol. A long-chain fatty acid is attached to the amino group of sphingosine through an amide linkage اللي قبل كان ( استر او في بعض ) الحاالت ايثر, producing a ceramide, which serve as a precursor of glycolipids. to مهمه: The alcohol group at carbon 1 of sphingosine is esterified phosphorylcholine, producing sphingomyelin, the only significant sphingophospholipid in humans. Sphingomyelin is an important componant of the myelin of nerve fibers (myelin sheath) that insulates and protects neuronal fibers of the central nervous system. Synthesis of Phospholipid 1. Glycerophospholipid (GP) synthesis involves : طرق التصنيع تتم بـاما either the1- donation of phosphatidic acid from CDP-diacylglycerol to an alcohol, or 2- the donation of the phosphomonoester of the alcohol from CDP-alcohol to 1,2-diacylglycerol. CDP is cytidine diphosphate. 2. In both cases, the CDP-bound structure is considered an "activated intermediate," and CMP is released as a side product of GP synthesis. 3. A key concept in phosphoglyceride synthesis, is activation either of diacylglycerol or the alcohol to be added by linkage with CDP. انظر الرسمه الصفحه القادمه تريحك من صداع الراس هذا الطريقه 1 في كل الحالتين يتم اخراج CMP الطريقه 2 الفرق بين الطريقتين فقط مكان وجود 4. The fatty acids esterifies to the glycerol alcohol groups can vary widely, contributing to the heterogeneity of this group of compounds. 5. Phospholipids are مهمهsynthesized in the Smooth ER-- then transported to Golgi apparatus and then membranes organelles or plasma membrane, or are secreted by exocytosis االخراج الخلوي. 6. There are two classes of phospholipids: Those have either glycerol as a backbone or contain sphingosine. كما شرح من قبل 7. Both classes are found in membranes and play a role in the generation of lipid-signaling molecules. A. Synthesis of phosphatidic acid (PA) 1. PA is the precursor الماده االوليهof many other phosphoglycerides. The steps in its synthesis from glycerol phosphate and two fatty acyl CoAs were in which PA is shown as a precursor of triacylglycerol. 2. All cells except mature erythrocytes can synthesize phospholipids, whereas triacylglycerol synthesis occurs only in liver, adipose tissue, lactating mammary glands, and intestinal mucosal cells. B: Synthesis of phosphatidylethanolamine (PE) & phosphatidylcholine (PC) • PC & PE are most abundant phospholipids in eukaryotic المخلوقات حقيقه النواهcells. • The primary route of synthesis uses choline and ethanolamine obtained either from the diet or turnover اعاده تصنيعof the body's phospholipids. • مهمهIn the liver, PC also can be synthesized from phosphatidylserine (PS) and PE. 1. Synthesis of PE and PC from preexisting choline and ethanolamine These synthetic pathways involve the phosphorylation of choline or ethanolamine by انزيمkinases, followed by conversion to the activated form, CDP-choline or CDP-ethanolamine ارجع الى الطريقه الثانيه لتصنيع الفسفولبدز لكي تتوضح. Finally, choline-phosphate or ethanolamine-phosphate is transferred from the nucleotide (leaving CMP) to a molecule of diacylglycerol (see Figure 17.5). Significance of choline reutilization اعاده استعمال: The reutilization of choline is important because, whereas humans can synthesize choline de novo, the amount made is insufficient for our needs. Thus, choline is an essential dietary nutrient with an adequate intake of 550 mg for men and 420 mg for women. Role of PC in lung surfactant 1. The principal pathway for the synthesis of dipalmitoylphosphatidylcholine 2 (داخله في التركيب االساسي للسيرفاكتينت انظر رقمDPPC, or dipalmitoylecithin). In DPPC, positions 1 and 2 on the glycerol are occupied by palmitate. 2. DPPC, made and secreted by granular pneumocytes, is the major lipid component of lung surfactant--the extracellular fluid layer lining the alveoli. 3. Surfactant serves to decrease the surface tension of this fluid layer, reducing the pressure needed to reinflate alveoli, thereby preventing alveolar collapse (atelectasis). 4. Respiratory distress syndrome (RDS) in pre-term infants is associated with insufficient surfactant production, and is a significant cause of all neonatal deaths in western countries. 5. Lung maturity of the fetus can be gauged by determining the ratio of DPPC to sphingomyelin, usually written as the L (for lecithin)/S ratio, in amniotic fluid. 6. A ratio of 2 or above ) النسبه السليمه ( مهمهis evidence دليلof maturity, because it reflects the major shift from sphingomyelin to DPPC synthesis that occurs in the pneumocytes at about 32 weeks of gestation. 7. ) مهمه )العالج1- Lung maturation can be accelerated تسرع by giving the mother glucocorticoids shortly before delivery. طريقه اخرى2-Administration of natural or synthetic surfactant (by intratracheal instillation تقطير عبر )القصبه الهوائيهis also used in the prevention and treatment of infant RDS. 8. Respiratory distress syndrome due to an insufficient amount of surfactant can also occur in adults whose surfactant producing pneumocytes have been damaged or destroyed, for example, as an adverse side effect of immuno-suppressive medication or chemotherapeutic drug use. 2: Synthesis of PC from phosphatidylserine (PS) in liver المكان مهم: 1. The liver requires a mechanism for producing PC, even when free choline levels are low, because it exports تصدرsignificant amounts of PC in the bile and as a component of serum lipoproteins. 2. To provide the needed PC, PS is decarboxylated تحول الىto phosphatidylethanolamine (PE) by بواسطه انزيمPS decarboxylase, an enzyme requiring pyridoxal phosphate as a cofactor. 3. PE then undergoes 3 methylation steps to produce PC, as illustrated in Figure 17.6. 4. S-adenosylmethionine (SAM) is the methyl group donor lead to S-adenosylhomocysteine C. Phosphatidylserine (PS) The primary pathway for synthesis of PS in mammalian tissues is provided by the base exchange reaction, in which the ethanolamine of PE is exchanged for free serine (see Figure 17.6). This reaction, although reversible, is used primarily to produce the PS required for membrane synthesis. Phospholipid Lectuer#2 D. Phosphatidylinositol (PI) PI is synthesized from free inositol and CDP-diacylglycerol as shown in Figure 17.5. الطريق االولى لصناعه الفسفولبدز ويمكن صناعته بالطريقه الثانيه كما في الشريحه القادمه مهمه: PI is an unusual phospholipid in that it often contains stearic acid on carbon 1 and arachidonic acid on carbon 2 of the glycerol. مهمه جداPI, therefore, serves as a reservoir خازنof arachidonic acid ( ) االركانيود اسد هو ماده اوليه تتحول الى عوامل االلتهاب: معلومهin membranes and, thus, provides the substrate for prostaglandin عامل التهاب synthesis when required. 1. Role of PI in signal transmission نقل االشارهacross membranes: a. The phosphorylation of membrane-bound phosphatidylinositol produces polyphosphoinositides, for example, phosphatidylinositol 4,5- bisphosphate (PIP2). b. The degradation of PIP2 by phospholipase C occurs in response to the binding of neurotransmitters, hormones, and growth factors to receptors on the cell membrane (Figure 17.8). c. The products of this degradation, inositol 1,4,5-trisphosphate (IP3) and diacylglycerol (DAG), mediate the mobilization of intracellular calcium and the activation of PKC, which act synergistically to evoke يستدعيspecific cellular responses. Signal transmission across the membrane is completed. نقل االشاره عبر الغشاء الخلوي -1يحدث فسفره للPI فينتج المركب اللي بالصوره -2عند وجود اشاره من الخارج كهرمون او غيره ينكسر المركب اللي بالصوره ويعطي الكحول ( اينوزيتول ) مع الفسفور و يعطي Diglycrol -3هذه العمليه تحفز االشارات الخلويه الصورة بالشريحه القادمه تسهل كل الموضوع مهمه وتسهل الموضوع وخصوصا االسهم 2. Role of PI in membrane protein anchoring تدعيم a. Specific proteins can be covalently برابطه تساهميهattached via a carbohydrate bridge to membrane bound PI (Figure 17.9). b. Examples of such proteins include alkaline phosphatase (a digestive enzyme found on the surface of the small intestine that attacks organic phosphates), and acetylcholine esterase (an enzyme of the postsynaptic membrane that degrades the neurotransmitter acetylcholine). c. Cell surface proteins bound to glycosyl phosphatidylinositol (GPI) are also found in a variety of parasitic protozoans ( الطفيلياتfor example, Trypanosomes and Leishmania). d. Being attached to a membrane lipid rather than being an integral part of the membrane) allows GPI-anchored proteins rapid lateral mobility on the surface of the plasma membrane. e. The protein can be cleaved from its anchor by the action of phospholipase C مهمهreleasing diacylglycerol. f. A deficiency in the synthesis of GPI in hematopoietic تكوين الدم cells results in a hemolytic disease, paroxysmal nocturnal hemoglobinuria. E. Phosphatidylglycerol (PG) and cardiolipin PG occurs in relatively large amounts in mitochondrial membranes and is a precursor of cardiolipin. It is synthesized by a two-step reaction from CDPdiacylglycerol and glycerol 3-phosphate. Cardiolipin (diphosphatidylglycerol, is composed of two molecules of phosphatidic acid connected by a molecule of glycerol. It is synthesized by the transfer of diacylglycerophosphate from CDP-diacylglycerol to a preexisting molecule of phosphatidylglycerol. F. Sphingomyelin Sphingomyelin, a sphingosine-based phospholipid, is a مهمهmajor structural lipid in the membranes of nerve tissue. The synthesis of sphingomyelin is shown in Figure, Briefly, palmitoyl CoA condenses with serine, as coenzyme A and the carboxyl group (as CO2) of serine are lost. This reaction, like the decarboxylation reactions involving amino acids, requires pyridoxal phosphate (a derivative of vitamin B6) as a coenzyme. The product is reduced in an NADPH-requiring reaction to sphinganine, which is acylated at the amino group with one of a variety of long-chain fatty acids, and then desaturated to produce مهمهa ceramide--the immediate precursor of sphingomyelin. مهمهA ceramide with a fatty acid thirty carbons long is a major component of skin, and regulates skin's water permeability. Phosphorylcholine from phosphatidylcholine is transferred to the ceramide, producing sphingomyelin and diacylglycerol. Sphingomyelin of the myelin sheath contains predominantly دائما longer-chain fatty acids such as lignoceric acid and nervonic acid. whereas gray matter of the brain has sphingomyelin that contains primarily stearic acid. IV. DEGRADATION OF PHOSPHOLIPIDS The degradation of phosphoglycerides is performed by phospholipases مهمه جدا جدا جداfound in all tissues and pancreatic juice. A number of toxins and venoms سمومhave phospholipase activity, and several pathogenic bacteria (Baccili) produce phospholipases that dissolve cell membranes and allow the spread of infection. ) مهمه ( حاله خاصهSphingomyelin is degraded by the lysosomal phospholipase, sphingomyelinase. A. Degradation of phosphoglycerides a. Phospholipases hydrolyze يكسرthe phosphodiester bonds of phospho- glycerides, with each enzyme cleaving phospholipid at a specific site. b. The major enzymes responsible for degrading phosphoglycerides are shown in Figure. c. Removal of the fatty acid from carbon 1 or 2 of a phosphoglyceride produces a lysophosphoglyceride, which is the substrate for lysophospholipases. d. Phospholipases release molecules that can serve as messengers (DAG and IP3), or that are the substrates for synthesis of messengers (arachidonic acid). e. مهمهPhospholipases are responsible not only for degrading phospholipids, but also for "remodeling "تغيير اشكالهمthem. f. For example, phospholipases A1 and A2 remove specific fatty acids from membrane-bound phospholipids; these can be replaced with alternative fatty acids using fatty acyl CoA transferase. g. This mechanism is used as one way to create the unique lung surfactant, dipalmitoylphosphatidylcholine (see p. 202), and to insure that carbon 2 of PI (and sometimes of PC) is bound to arachidonic acid. B. Degradation of sphingomyelin a. يتم الفصل على مرحلتين1-Sphingomyelin is degraded by sphingomyelinase, a lysosomal enzyme that hydrolytically removes phosphorylcholine, leaving a ceramide مهمه. 2-The ceramide is, in turn, cleaved by ceramidase into sphingosine and a free fatty acid (Figure 17.12). b. The ceramide and sphingosine released by the degradation of sphingomyelin play a role as intracellular messengers. c. مهممهCeramides appear to be involved In the response to stress, and sphingosine inhibits protein kinase C. d. مهمهNiemann-Pick disease (Types A and B) is an autosomal recessive disease caused by the inability to degrade sphingomyelin. The deficient enzyme is sphingomyelinase-A type of phospholipase C. e. In the severe infantile form (type A), the liver and spleen are the primary sites of lipid deposits and are, therefore, tremendously enlarged. The lipid consists primarily of the sphingomyelin that cannot be degraded (Figure 17.13). f. Infants with this disease have rapid, progressive neurodegeneration deposition of sphingomyelin in CNS, and they die in early childhood. g. A less severe variant (type B) causes little to no damage to neural tissue, but lungs, spleen, liver, and bone marrow are affected, resulting in a chronic form of the disease, with a life expectancy only to early adulthood. h. Although Niemann-Pick disease occurs in all ethnic groups. both type A and B occur with greater frequency in the Ashkenazi Jewish ) يهود اشناكاز ( احسسسسسسسسسسسسن population than in the general population. i. In the Ashkenazi Jewish population, the incidence of type A is 1:40,000 live births, and that of type B is 1:80,000. j. The incidence of Niemann-Pick disease in the general population is less than 1:100,000. Conclusions 1. Structure of Glycerophospholipids and sphingolipids. 3. Synthesis of phospholipids: Phosphatidic acid, PC, PE and PC, cardiolipin and sphingomyelin etc. 4. Degradation of phospholipids by phosphalipases. 5. Role of phosphatidyl choline in lung surfactant 6. Role of PI in signal transduction and membrane anchoring . 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