Biochemistry 7 - Metabolism III PDF

Amino Acids oxidation (the urea cycle) Amino acids The amino acids have an amino and a carboxylic acid group that are joined to the same carbon atom. This carbon atom is referred to as the α carbon. AAs...

Amino Acids oxidation (the urea cycle) Amino acids The amino acids have an amino and a carboxylic acid group that are joined to the same carbon atom. This carbon atom is referred to as the α carbon. AAs differ from each other in their side chains, or R groups, which vary in structure, size, and electric charge, and which influence the solubility of the amino acids in water Key principles The many paths for AA catabolism have two broad parts: one involving the amino groups the other involves the carbon skeletons Four AAs – alanine, glutamate, glutamine, and aspartate play key roles in the transport and distribution of amino groups Free ammonia is toxic Each amino acid has a different catabolic fate Metabolic circumstances of AA oxidation AA undergo oxidative degradation when: AAs released during protein turnover are not needed for new protein synthesis Ingested AA exceed the body’s needs for protein synthesis Cellular proteins are used as fuel because carbs are either unavailable or not properly utilized Protein turnover Daily protein turnover for humans is about 300g AAs contain nitrogen atoms, which need to be eliminated without developing too much toxic ammonia Steps of the urea cycle occur in the liver Described in 1932 – first described cyclic metabolic pathway (Hans Krebs and Kurt Henseleit, 1932), five years before the discovery of the TCA cycle https://www.bioc.cam.ac.uk/about-us/history/nobel-prizes/hans-adolf-krebs Metabolic Fates of Amino groups Unless reused amino groups are channeled into a single excretory end product – urea Glutamate, glutamine, alanine, and aspartate are most easily converted into citric acid cycle intermediates: Glutamate and glutamine to alpha-ketoglutarate Alanine to pyruvate Aspartate to oxaloacetate TCA cycle accepts carbon skeletons The cycle accepts 3-, 4-, 5- carbon skeletons The breakdown of amino acids yields carbon skeletons: Deaminated aspartate yields oxaloacetate Deaminated glutamate yields alpha-ketoglutareate Protein metabolism and Urea cycle During digestion, proteins are hydrolysed into amino acids Amino acids are oxidized via Krebs cycle after conversion by processes such as deamination, decarboxylation or hydrogenation GOAL – eliminate ammonia from the body by converting it to urea  Ammonia originates in the catabolism of amino acids that are primarily produced by the degradation of proteins – dietary as well as existing within the cell Digestive enzymes Proteins released by digestion of cells Muscle proteins Haemoglobin Intracellular proteins (damaged, unnecessary) Ammonia is toxic, especially for the CNS, because it reacts with alpha-ketoglutarate to form glutamate, thus making it limiting for the Krebs cycle – decrease in the ATP level Also it competes with K+ for transport into astrocyte cells viaNa+K+ATPase – resulting in elevated extracellular K+ (follows excess Cl- which alters neuronal response to the neurotransmitter GABA) Liver damage or metabolic disorders associated with elevated ammonia can lead to tremor, slurred speech, blurred vision, command and death  2 CHOICES 1. Reuse 2. Urea cycle = organic compounds that contain a carboxylic acid group (−COOH) and a Fumarate ketone group (>C=O) Oxaloacetate UREA CYCLE Ammonia is converted into urea in the liver (mitochondria of hepatocytes) and excreted in urine The carbon and oxygen of urea are derived from CO2 Urea is produced by the liver, and then is transported in the blood to the kidneys for excretion in the urine  Carboxylic acid oxidation Waste or reuse Amino acids catabolism Urea Ammonia Nitrogen removal from amino acids Step 1: Remove amino group Step 2: Take amino group to liver for nitrogen excretion Step 3: Entry into mitochondria Step 4: Prepare nitrogen to enter urea cycle Step 5: Urea cycle  Nitrogen removal from amino acids 2 ways: Pyridoxal phosphate PLP Transamination – transfer of the AA Aminotransferase - Cofactor (coenzyme) Transamination group to alpha-keto acid, which is PLP - Active from of vit B6 then converted to AA Deamination – release of the AA Transamination – in the skeletal muscle group as ammonia NH3 cells and then delivered into the Oxidative bloodstream and the liver (citric acid deamination cycle) Urea cycle Step 1. Remove amino group Transfer of the amino group of an amino acid to an -keto acid  the original AA is converted to the corresponding -keto acid and vice versa:  Transamination is catalyzed by transaminases (aminotransferases) that require participation of pyridoxalphosphate: Used as a prosthetic group by all aminotransferases Carries amino groups at the active site amino acid pyridoxalphosphate Schiff base Step 2: Take amino group to liver for nitrogen excretion Glutamate releases its amino group as ammonia in the liver The amino groups from many of the Glutamate -amino acids are collected in the dehydrogenase liver in the form of the amino group of L-glutamate molecules The glutamate dehydrogenase of mammalian liver has the unusual capacity to use either NAD+ or NADP+ as cofactor Nitrogen carriers 1. Glutamate transfers one amino group WITHIN cells: Aminotransferase → makes glutamate from -ketoglutarate Glutamate dehydrogenase → opposite 2. Glutamine transfers two amino groups BETWEEN cells → releases its amino group in the liver 3. Alanine transfers amino groups from tissue (muscle) into the liver alanine aminotransferase – interconverts pyruvate and alanine via transamination with glutamate Skeletal muscles produce pyruvate, lactate and ammonia  Glucose-alanine cycle (Cahil cycle) Alanine plays a special role in transporting amino groups to liver. Alanine is the carrier of ammonia and of the carbon skeleton of pyruvate from muscle to liver. The ammonia is excreted and the pyruvate is used to produce glucose, which is returned to the muscle. Cori cycle? According to D. L. Nelson, M. M. Cox :LEHNINGER. PRINCIPLES OF BIOCHEMISTRY Fifth edition Step 3: entry of nitrogen to mitochondria Step 4: prepare nitrogen to enter urea cycle Regulation Reaction steps of the urea cycle The first two reactions leading to the synthesis of urea occur in the mitochondria, whereas the remaining cycle enzymes are located in the cytosol Formation of carbamoyl phosphate (M) Formation of citrulline (M) Formation of argininosuccinate Cleavage of argininosuccinate to arginine and fumarate Hydrolysis of arginine Step 5: Urea cycle aspartate Ornithine transcarbamoylase Argininosuccinate synthase Arginase 1 Argininosuccinate lyase Urea cycle – review (Sequence of reactions) Carbamoyl phosphate formation in mitochondria is a prerequisite for the urea cycle (Carbamoyl phosphate synthetase I) Citrulline formation from carbamoyl phosphate and ornithine (Ornithine transcarbamoylase) Aspartate provides the additional nitrogen to form argininosuccinate in cytosol (Argininosuccinate synthase) Arginine and fumarate formation (Argininosuccinate lyase) Hydrolysis of arginine to urea and ornithine (Arginase) UREA CYCLE RECAP in pics Regulation of urea cycle The activity of urea cycle is regulated at two levels: Dietary intake is primarily proteins  urea (amino acids are used for fuel) Prolonged starvation  breaks down of muscle proteins  urea also The rate of synthesis of four urea cycle enzymes and carbamoyl phosphate synthetase I (CPS-I) in the liver is regulated by changes in demand for urea cycle activity Enzymes are synthesized at higher rates in animals during: starvation in very-high-protein diet Enzymes are synthesized at lower rates in well-fed animals with carbohydrate and fat diet animals with protein-free diets Regulation of urea cycle N-acetylglutamic acid – allosteric activator of CPS-I High concentration of Arg → stimulation of N-acetylation of glutamate by acetyl-CoA  Essential and Non essential AA Phenylketonuria – mutation in the enzyme phenylalanine hydroxylase https://aminoco.com/blogs/amino-acids/conditionally-essential-amino-acids Lipids - review Lipids are a heterogeneous group of compounds, including fats, oils, steroids, waxes, and related compounds, which are related more by their physical than by their chemical properties: insolubility in water and solubility in nonpolar solvents Lipids are important in biological systems: they form the cell membrane, provide energy for life and several essential vitamins are lipids Three major subclasses are recognised: simple, compounds and steroids Lipids can be also divided in two major classes: nonsaponifiable lipids and saponifiable lipids Classification Again there are different means of classifying the steroids Here, I present one based on the type of substituent group at C-17, i.e., group R 1. Sterols – R is an aliphatic side chain, containing usually one or more hydroxyl groups 2. Sex hormones – R bears a ketonic or hydroxyl group and mostly possess a two carbon side chain 3. Cardiac glycoside – R is a lactone ring (contains sugar) 4. Bile acids – R is essentially a five-carbon side chain ending with a carboxylic acid 5. Sapongenins – R contains an oxacyclic ring system 3 Stages of Fatty acid oxidation Stage 1: beta-oxidation - Fatty acids undergo oxidative removal of successive two-carbon units in the form of acetyl-CoA Stage 2: oxidation of acetyl-CoA groups to CO2 in the citric acid cycle - Occurs in the mitochondrial matrix - Generates NADH, FADH2, and one GTP Stage 3: electron transfer chain and oxidative phosphorylation - Generates ATP from NAD and FADH2 Beta oxidation Beta-oxidation is the catabolic process by which fatty acid molecules are broken down in the mitochondria in eukaryotes to generate acetyl-CoA Acetyl-CoA enters the citric acid cycle while NADH and FADH2, which are co-enzymes, are used in the electron transport chain Or the formation of keton bodies.. Substrates: Free fatty acids; H2O Products: One acetyl CoA, one NADH, and one FADH2 for every removal of a two-carbon group from the fatty acid chain It is referred as “beta-oxidation” because the beta carbon of the fatty acid undergoes oxidation to a carbonyl group  Carboxyl group β Overview of the beta oxidation process blood Adipose tissue Triglycerides Fatty acids Cytoplasm of the cell Matrix of mitochondria Oxidative metabolism Beta oxidation A) Transport from adipose tissue to target cells B) Entry into cytoplasm and mitochondria C) Oxidative catabolism inside mitochondrial matrix Fatty Acids are preferentially oxidized: During periods of extended exercise e.g. aerobics, running on a treadmill, running for long distances In diabetic patients in whom glucose metabolism is low During periods of starvation By heart muscle which almost exclusively depends on FA oxidation for energy A) Transport from adipose tissue to target cells Lipase Free fatty acid Lack mitochondria  B) Entry into cytoplasm and mitochondria FA in Plasma FA Bound to FABP Negative (transporter) charge, cannot cross as such porins  Carnitine 1. shuttle 2. 3. C) Oxidative catabolism inside mitochondrial matrix FAO Cycle Fatty acid (14, 16..C length) 4 enzyme steps: Dehydrogenation (2x) Hydration Thiolysis (cleavage) Energy Yield for Even-chain Fatty Acids Energy is generated from the products of β-oxidation The 16-carbon palmitoyl-CoA undergoes seven repetitions In the last repetition, a 4-carbon fatty acyl-CoA (butyryl-CoA) is cleaved to two acetyl-CoAs 1. When one palmitoyl-CoA is oxidized, seven FADH2, seven NADH, and eight acetyl-CoA are formed. The seven FADH2 each generate approximately 1.5 ATP, for a total of about 10.5 ATP. The seven NADH each generate about 2.5 ATP, for a total of about 17.5 ATP. The eight acetyl-CoA can enter the TCA cycle, each producing about 10 ATP, for a total of about 80 ATP. From the oxidation of palmitoyl-CoA to CO2 and H2O, a total of about 108 ATP are produced. 2. The net ATP produced from palmitate that enters the cell from the blood is about 106 because palmitate must undergo activation (a process that requires the equivalent of 2 ATP) before it can be oxidized (108 ATP − 2 ATP = 106 ATP). 3. The oxidation of other fatty acids will yield different amounts of ATP Oxidation of odd-chain and unsaturated fatty acids Odd-chain fatty acids produce acetyl-CoA and propionyl-CoA Propionyl CoA can be converted to succinyl CoA through three enzymatic events, which require biotin and vitamin B12 as cofactors, and then succinyl CoA can enter the citric acid cycle Unsaturated fatty acids, which comprise about half the fatty acid residues in human lipids, require enzymes in addition to the four that catalyze the repetitive steps of the β-oxidation spiral  FA synthesis A process by which the saturated FA (palmitate – C16) is formed Excess carbohydrates are converted to FA, creating a store of energy able to be broken down for future energy Regulation of FAO 1. Enzyme CPTI (carnitine-palmitoyl transferase I) is the rate-limiting enzyme. It is inhibited by Malonyl CoA, a product formed during fatty acid synthesis 2. Hormonal Regulation of FA oxidation Glucagon Epinephrine Insulin Ketone bodies Ketone bodies – acetoacetate => acetone and hydroxybutyrate Formed from acetyl-CoA in the liver Acetone is exhaled Acetoacetate and hydroxybutyrate are transported to extrahepatic tissues and converted to acetyl CoA to be oxidized in the citric acid cycle Ketone bodies are used as fuels in all tissues except the liver The liver lacks an enzyme (beta-ketoacyl-CoA transferase) The liver is a producer of ketone bodies Overproduced in diabetes and during starvation