Metabolism of Glycolipids, Isoprenoids & Steroids Lecture 9 PDF

Metabolism of glycolipids, isoprenoids and steroids Dr. Ardina Grüber Nanyang Technological University School of Biological Sciences Division of Structural Biology and Biochemistry...

Metabolism of glycolipids, isoprenoids and steroids Dr. Ardina Grüber Nanyang Technological University School of Biological Sciences Division of Structural Biology and Biochemistry Singapore 637551 email: [email protected] According to their chemical structure lipids are organized in some classes of lipids: Marks Basic Medical Biochemistry: A clinical Approach, 4th edition  Glycerol The phospholipids, which include both glycerophospholipids and sphingomyelins, are crucial components of membrane structure. They are also precursors of hormones such as eicosanoids and signal molecules. Voet, Voet: BIOCHEMISTRY 3rd edition  Pathways in glycerophospholipid biosynthesis The major phospholipids found in membranes are shown in green. Pathways found in both bacterial and eukaryotic cells are highlighted in light purple. Other reactions are confined to eukaryotic cells. DHAP = dihydroxyacetone phosphate; DAG = diacylglycerol; AdoMet = S-adenosylmethionine  The synthesis of glycerolipids in eukaryotes begins with the formation of phosphatidic acid, which may be formed from glycerol, diacylglycerol or dihydroxyaceton phosphate. Glycerokinase catalyzes the phosphorylation of glycerol to form glycerol-3-phosphate, which is then acylated at both the 1- and 2-positions to yield phosphatic acid. Diacylglycerol kinase ATP ADP P H20 Phosphatidic acid phosphatase Garrett, Grisham: Biochemistry 4th edition Eukaryotic systems can also utilize dihydroxyaceton phosphate as a starting point for synthesis of phosphatidic acid. Alternatively, dihydroxyacetone phosphate can be reduced to glycerol-3-phosphate by glycerol-3-phosphate dehydrogenase. Garrett, Grisham: Biochemistry 4th edition  In eukaryotes, phophatidic acid is converted directly either to diacylglycerol or to cytidine diphosphodiacylglycerol (CDP- diacylglycerol). Diacylglycerol is a precursor for synthesis of triacylglycerol, phosphatidylethanolamine, and phosphatidylcholine. Triacylglycerol is synthesized mainly in adipose tissue, liver, and intestines and serves as the principal energy storage molecule. Its biosynthesis in liver and adipose tissues occurs via diacylglycerol acyltransferase, an enzyme bound to the cytoplasmic face of the ER. Garrett, Grisham: Biochemistry 4th edition The biosynthesis of triacylglycerols is different in intestines. Recall that triacylglycerols from the diet are broken down to 2-monoacylglycerols by specific lipases. Acyltransferases then acylate 2-monoacylglycerol to produce new triacylglycerols. Garrett, Grisham: Biochemistry 4th edition Garrett, Grisham: Biochemistry 4th edition  Mammals synthesize phosphatidylserine (PS) in a calcium ion-dependent reaction involving aminoalcohol exchange. The enzyme catalyzing this reaction is associated with the ER and will accept phosphatidylethanolamine (PE) and other phospholipid substrates. A mitochondrial PS decarboxylase can subsequently convert PS to PE. Garrett, Grisham: Biochemistry 4th edition  CDP-diacylglycerol as precursor of glycerolipids in eukaryotes In eukaryotes, phophatidic acid is converted directly either to diacylglycerol or to cytidine diphosphodiacylglycerol (CDP-diacylglycerol). From these two precursors, all other glycero- phospholipids in eukaryotes are derived. Garrett, Grisham: Biochemistry 4th edition  Phosphatidic acid is activated as CDP-diacylglycerol The reaction of CTP with phosphatidic acid is catalyzed by CDP-diacylglycerol synthase (reaction 1). This reaction is drawn to the right by the enzymatic hydrolysis of pyrophosphate, catalyzed by the ubiquitous pyrophosphatase (reaction 2). CDP-diacylglycerol is activated for nucleophilic attack by various polar head groups (CMP is a good leaving group). © 2016 Pearson Education, Ltd.  Eukaryotes also use CDP-diacylglycerol, derived from phosphatidic acid, as a precursor for several other important phospholipids, including phosphatidylinositol (PI), phosphatidylglycerol (PG), and cardiolipin. Garrett, Grisham: Biochemistry 4th edition  β-ketosphinganine synthase (PLP) Condensation of palmitoyl-CoA and serine followed by reduction with NADPH yields sphinganine, which is then β-ketosphinganine acylated to N-acylsphinganine. reductase Mixed function Oxidase (animals) Lehninger: Biochemistry 4th edition  Ceramide is the building block for all other sphingolipids. Sphingomyelin, for example, is produced by transfer of phosphocholine from phohatidylcholine. Glycosylation of ceramide by sugar nucleotides yields cerebrosides, such as galactosylceramide, which make up 15% of the lipids of myelin sheath structures. Globosides- more than one sugar. Garrett, Grisham: Biochemistry 4th edition  Cerebrosides that contain one or more sialic acid (N-acetylneuraminic acid) moieties are called gangliosides. Several dozen gangliosides have been characterized, and the general form of the biosynthetic pathway is illustrated for the case of ganglioside GM2. Garrett, Grisham: Biochemistry 4th edition  locally acting signaling molecules Eicosanoids, so named because they are derived from C20 fatty acids, are breakdown products of phospholipids. Biologically active eicosanoids are short- lived, locally acting hormones. In humans, the most prevalent precursor of eicosanoids is a arachidonic acid, a C-20 polyunsaturated fatty acid that has four nonconjugated double bonds. cis - 5, 8, 11, 14-eicosatetraenoic acid or Arachidonic acid Voet, Voet: BIOCHEMISTRY 3rd edition  Prostaglandins, thromboxanes and leukotrienes are important eicosanoids synthesized from arachidonic acids Arachidonic acid is the precursor of prostaglandins (PG), thromboxanes (Tx), and leukotriene. Thromboxanes were discovered in thrombocytes. Leukotriene C was originally discovered in the class of white blood cells called polymorphonuclear leukocytes and was named after the source (leukocytes) and the triene structure (three conjugated double bonds). The key step in the synthesis of prostaglandins (involved in inflammation) is PGH synthase, a dual function enzyme. Aspirin inhibits the COX-1 and COX-2 forms of PGH synthase, yielding its anti- ©inflammatory activity. 2016 Pearson Education, Ltd.  PLA2: phosphatidylcholine phosphatidylethanolamine PLC: phosphaditylinositol Voet, Voet: BIOCHEMISTRY 3rd edition  All prostaglandins are cyclopentanoic acids derived from arachidonic acid. COX The biosynthesis of prostaglandins is initiated by an enzyme associated with the ER, called prostaglandin endoperoxide H synthase (PGHS), also known as cyclooxygenase (COX). The enzyme that converts arachidonic acid to prostaglandin PGH2, possesses two distinct activities: cyclooxygenase (COX) and a gluthatione-dependent hydroxyperoxidase (POX). POX Voet, Voet: BIOCHEMISTRY 3rd edition Prostaglandin H2 which is derived from arachidonic acid, is a precursor for many other biologically significant molecules. X-Ray structure of PGH synthase (PGHS) from sheep. Inactivation of PGH synthase by aspirin. Heme, fluriprofen, and Tyr 385 are labeled. Aspirin (acetylsalicylate) exerts most of its effects by inhibiting the biosynthesis of prostaglandins. Its site of action is PGHS. COX activity is destroyed when aspirin O-acetylates Ser530 on the enzyme. Aspirin exerts its powerful anti-inflammatory effect by inhibiting the first step in their synthesis. Garrett, Grisham: Biochemistry 4th edition  Science 2007: vol. 318 pp. 1258-1265 Cholesterol The basic ring structure of sterols; the perhydrocyclopentanophenanthrene nucleus. Steroids constitute a class of lipids that are classified as isoprenoids because their structures are related to the five-carbon molecule isoprene. Steroids contain four fused rings: three six-carbon rings designated as A, B, C and five-carbon D ring. Cholesterol plays an essential role in mammalian biochemistry. It is not only a component of certain membranes but is also a precursor of steroid hormones and bile salts. Mathews, van Holde, Ahern: Biochemistry 3rd edition Cholesterol biosynthesis can be divided into three distinct processes 1. Conversion of C2 fragments (acetate) to a C6 isoprenoid precursor (mevalonate) 2. Conversion of six C6 mevalonates, via activated C5 intermediates, to the C30 squalene 3. Cyclization of squalene and its transformation to the C27 cholesterol 3 Voet, Voet: BIOCHEMISTRY 3rd edition  The cholesterol biosynthesis pathway begins in the cytosol with the synthesis of mevalonate from Acetyl-CoA. The rate-limiting step in cholesterol biosynthesis is catalyzed by the HMG-CoA reductase, responsible for the formation of 3R-mevalonate from 3-Hydroxy-3-methylglutaryl-CoA. Here, HMG-CoA undergoes two NADPH- dependent reductions to produce 3R-mevalonate. Garrett, Grisham: Biochemistry 4th edition The biosynthesis of squalene involves conversion of mevalonate to two key 5- c a r b o n i n t e r m e d i a t e s , i s o p e n t e ny l py ro p h o s p h a te a nd d i m e t hy la l ly l pyrophosphate (next slide), which joins to yield farnesyl pyrophosphate and then squalene. Garrett, Grisham: Biochemistry 4th edition  Isomerization of the double bond in isopentenyl pyrophosphate yields the dimethylallyl pyrophosphate. Condensation of these two 5-carbon intermediates produces geranyl Occurs in the cytosol pyrophosphate; addition of another 5- carbon isopentenyl group gives farnesyl Geranyl pyrophosphate pyrophosphate. Both steps in the production of farnesyl pyrophosphate occur with release of pyrophosphate, hydrolysis of which drives these reactions forward. Squalene synthase (farnesyl transferase) is bound to the membrane of the ER. Occurs in the ER Garrett, Grisham: Biochemistry 4th edition  Squalene monooxygenase, an enzyme bound to the ER, converts squalene to squalene-2,3-epoxide. This reaction employs FAD and NADPH as coenzymes and requires O2. A second ER membrane enzyme, 2,3-oxidosqualene lanosterol cyclase, catalyzes the second reaction, which involves a succession of 1,2 shifts of hydride ions and methyl groups. Garrett, Grisham: Biochemistry 4th edition  Although lanosterol may appear similar to cholesterol in structure, another 20 steps are required to convert lanosterol to cholesterol. The enzymes responsible for this are all associated with the ER. Garrett, Grisham: Biochemistry 4th edition  Cholesterol has several fates A small fraction of the cholesterol made in liver is incorporated into the membranes of hepatocytes, but most of it is exported in one of three forms: biliary cholesterol, bile acids and cholesteryl esters, Cholesterol will be used for the synthesis of steroid hormone and as a precursor of vitamin D.  An α-hydroxyl group is formed at position 7 of cholesterol. This reaction, which is inhibited by bile salts, is the rate-limiting step in bile salt synthesis. Marks Basic Medical Biochemistry: A clinical Approach, 4th edition  Nascent HDL is synthesized in liver and intestinal cells. It exchanges proteins with chylomicrons and VLDL. HDL picks up cholesterol (C) from cell membranes. This cholesterol is converted to cholesterol ester (CE) by the enzyme lecithin-cholesterol acyltransferase (LCAT) dependent reaction. HDL transfers CE to VLDL in exchange for triacylglycerol (TG). The cholesterol ester transfer protein (CETP) mediates this exchange.  Overview of LDL receptors in cholesterol uptake and metabolism © 2016 Pearson Education, Ltd.  Desmolase The biosynthesis of steroid hormones begins with the desmolase reaction, which converts cholesterol to pregnenolone. Desmolase is found in the mitochondria of tissues that synthesize steroids. Pregnenolone is transported from the mitochondria to the ER, where a hydroxyl oxidation and migration of the double bond yield progesterone. Pregnenolone synthesis in the adrenal cortex is activated by adrenocorticotropic hormone (ACTH), a peptide of 39 amino acid residues secreted by the anterior pituitary gland. Progesterone is also the precursor for synthesis of the other sex hormone steroids and the corticosteroids. Voet, Voet: BIOCHEMISTRY 3rd edition Regulation of cholesterol synthesis Rate-limiting step is conversion of HMG CoA to mevalonate. Cholesterol synthesis is regulated by: HMG-CoA reductase is regulated covalently by PP2A, PKA, and by AMPK Glucagon (promotes phosphorylation - inactivation of HMG-CoA reductase;) Insulin (promotes dephosphorylation- activation of HMG-CoA reductase). Intracellular cholesterol concentrations X represents metabolites of cholesterol that stimulate proteolysis of HMG-CoA reductase (oxysterol).  To Make a Cholesterol (To the tune of "When Johnny Comes Marching Home") Copyright © 2010 Kevin Ahern Some things that you can build with acetyl-CoAs Are joined together partly thanks to thiolase They come together 1-2-3 Six carbons known as H-M-G And you’re on your way To make a cholesterol To synthesize a mevalonate in the cell Requires reducing HMG-CoA, as well The enzyme is a RE-ductase Controlled in allosteric ways When the cell's impelled To make a cholesterol. The mevalonate made in metabolic schemes Gets decarboxylated down to isoprenes They’re linked together willy-nil To build a PP-geranyl In the cells’ routines To make a cholesterol A single step links farnesyls but that’s not all The squalene rearranges to lanosterol From that there’s nineteen steps to go Before the sterol’s apropos Which you must recall To make a cholesterol The regulation of the scheme’s complex in ways Inhibited by feedback of the RE-duc-tase And statins mimic so they say The look of HMG-CoA So we sing their praise And not make cholesterol