* Your assessment is very important for improving the workof artificial intelligence, which forms the content of this project
Download Amino Acids Objectives
Basal metabolic rate wikipedia , lookup
Clinical neurochemistry wikipedia , lookup
Oligonucleotide synthesis wikipedia , lookup
Oxidative phosphorylation wikipedia , lookup
Nucleic acid analogue wikipedia , lookup
Ribosomally synthesized and post-translationally modified peptides wikipedia , lookup
Artificial gene synthesis wikipedia , lookup
Catalytic triad wikipedia , lookup
Point mutation wikipedia , lookup
Nitrogen cycle wikipedia , lookup
Metalloprotein wikipedia , lookup
Fatty acid synthesis wikipedia , lookup
Glyceroneogenesis wikipedia , lookup
Proteolysis wikipedia , lookup
Peptide synthesis wikipedia , lookup
Fatty acid metabolism wikipedia , lookup
Protein structure prediction wikipedia , lookup
Genetic code wikipedia , lookup
Citric acid cycle wikipedia , lookup
Biochemistry wikipedia , lookup
Amino Acids: Parts 1 and 2 1. Which amino acids are the essential and which are nonessential? Which nonessential amino acids are derived directly from an essential amino acid? Which amino acids are conditionally essential and why? Cysteine is made from methionine. If tyrosine is not adequate in the diet, large amounts of phenylalanine are required in order to produce it. Arginine and histidine are not synthesized in adequate quantities only when they are supporting growth in young animals. 2. How are amino acids absorbed in small intestine? Dietary protein is denatured under acidic conditions and partially digested by pepsin in the stomach. Broken down in the small intestine by trypsin, chymotrypsin, elastase, aminopeptidases, and carboxypeptideases. Didpeptidase (enterocytes) digests proteins and peptides. Amino acids are absorbed by these enterocytes on the luminal side by multiple carriers. Dipeptides use H+-dependent carriers. Separate basolateral carriers transport amino acids into the blood. Types of carriers that bring amino acids into the enterocytes: 1.)Na+-dependent amino acid transport systems with overlapping amino acid specificities 2.)Na+ and amino acids at high luminal concentrations are co-transported due to a concentration gradient into cells 3.)The ATP-dependent Na+/K+ pump exchanges the intracellular Na+ for extracellular K+, reducing Na+ levels to maintain the extracellular Na+ concentration gradient to drive transport 4.)Small peptides are transported by a proton (H+) driven transporter and hydrolyzed by intracellular peptidases. 5.)Amino acids in blood and extracellular fluids are transported by at least 7 ATPrequiring active transport systems with overlapping amino acid specificities 3. What molecules serve as carriers of nitrogen? Urea – ureotelic organisms; mammals. Nontoxic, highly water soluble (up to 10M), used only for excretion of excess nitrogen. Formed from ammonia and bicarbonate. Tranfers nitrogen in blood for excretion in the urine. Purines – purinotelic organisms; bats, birds, reptiles. Excreted in feces. Minimizes use of water. Great fertilizer. Ammonia – most invertebrates. Relatively small amounts excreted normally in urine by humans. Fish excrete ammonia through their gills. Small organisms can excrete ammonia directly into their environment. Glutamine – relatively nontoxic. Formed from glutamate and ammonia. Water soluble. An amino group donor in a number of pathways (synthesis of a lot of purines). Carrier of nitrogen to liver from muscles for synthesis of urea. Glu = Glutamate, Gln = Glutamine Alanine – nontoxic. Formed from pyruvate and an amino acid. Used for inter-organ transfer of nitrogen (glucose-alanine cycle). 4. What properties of urea make it well suited as a carrier of nitrogen? It’s nontoxic, highly water soluble, and used only for the excretion of nitrogen (a “metabolic dead end”). 5. Explain how transaminases and glutamate dehydrogenase play a central role in nitrogen metabolism. Transaminases transfer amino groups from amino acids (mainly alanine and glutamine) to α-ketoglutarate to make glutamate. Glutamate dehydrogenase then uses NAD+ to release ammonia for urea synthesis and be regenerated to α-ketoglutarate (oxidative deamination of glutamate). Glutamate dehydrogenase can use NADP+, but the predominant form used in vivo is NADPH toward glutamate and NAD+ toward αketoglutarate. It is confined to the mitochondrial matrix. 6. Under what conditions are ammonium ions excreted in relatively high concentrations in the urine? What enzyme plays an important role under these conditions? Metabolic acidosis. Glutaminase plays an important role under these conditions. (Note: urea production is suppressed. Nitrogen is being directly excreted as NH4+.) Glutamine + H2O → Glutamate + NH4+ 7. Explain how the glucose alanine cycle operates in inter-organ metabolism. Why is this cycle important? It’s important to provide for a system to transport nitrogen generated by metabolism of amino acids from one organ to another in a nontoxic form. It also produces pyruvate for production of glucose by gluconeogenesis in the liver. If you start in muscle, pyruvate (from metabolism of glucose) can be converted to alanine by transferring an amino group from some other amino acid to pyruvate by alanine aminotransferase (glutamate pyruvate amino transferase, GPT). This can be transferred in the blood to the liver, where it’s either directly deaminated to give ammonia, or it’s transferred to another amino acid, which goes to glutamate, which is eventually released as ammonia. One nitrogen from alanine is recovered to produce ammonia, which goes to production of urea. What is left from alanine produces pyruvate. Pyruvate is a gluconeogenic precursor, so glucose can be formed. Liver Muscle 8. Explain how pyridoxal phosphate functions in amino group transfer by transaminases. Pyridoxal phosphate is an electrophile for covalent catalysis in amino group transfer by transaminases. 9. How are the reactions of the urea cycle compartmentalized inside cells? In the mitochondria – First: activated 5-10 fold by N-acetylglutamate Second: Ornithine is made in the cytosol. Ornithine and citrulline require a membrane carrier to enter or exit the mitochondria Citrulline is then transported to the cytosol where the next three reactions occur. In the cytosol – Third: Citrulline is used to make argininosuccinate Fourth: Arginine and fumarate are formed from arginosuccinate Finally: Arginase releases urea from arginine. Arginase is present in the liver, but absent in the kidney (which is a major site of arginine synthesis). !!SUMMARY!! 10. Explain the role of the shuttles for ornithine/citrulline and malate/aspartate in the urea cycle. The malate/aspartate shuttle moves aspartate, glutamate, and α-ketoglutarate across the mitochondrial membrane by converting malate to oxaloacetate, and that to aspartate. Malate can be transported into the mitochondria as an antiport with αketoglutarate. Aspartate can be transported into the cytosol as an antiport with glutamate. Ornithine/citrulline is an antiport that carries citrulline and ornithine across the mitochondrial membrane. This is important because citrulline is being produced in the mitochondria during the second step of the urea cycle but reacts in the third step in the cytosol. Ornithine, on the other hand, is one of the final products of the urea acid cycle and can be transported back to the mitochondria to function as a reactant in the second reaction to make more citrulline. (See above image.) 11. How are the urea cycle and the TCA cycle linked? The urea cycle, through arginosuccinate lyase, produces fumarate. Fumarate can be combined with water via fumarase to produce malate, which is an intermediate in the TCA cycle. Aspartate can give rise to oxaloacetate through the action of a transaminase. So you can get aspartate from oxaloacetate in the TCA cycle, and then the aspartate can be used in the urea cycle to form arginosuccinate. 12. How is the urea cycle regulated? Long term regulation – elevated synthesis of urea cycle enzymes due to increased metabolism of amino acids, either from diet or from protein turnover (starvation). If in the case of a high protein diet, the carbon skeletons derived from amino acids in the diet are oxidized or converted to fat or glycogen. The nitrogen is eliminated through the synthesis of urea. Enzymes of the urea cycle are induced up to 20 fold over a low protein diet. In the case of starvation, the carbon skeletons derived from the breaking down of muscle (protein catabolism) are oxidized and nitrogen is used for urea. Enzymes of urea cycle are induced. Short term regulation – Carbamoyl phosphate synthetase I is allosterically activated 5-10 fold by N-acetylglutamate. Synthesis of N-acetylglutamate depends on the availability of Acetyl-CoA and glutamate. Arginine activates N-acetylglutamate synthase. The precursors to N-acetylglutamate and arginine are signals of high levels of amino acids whose metabolism will generate ammonia. Acetyl-CoA + glutamate → N-acetylglutamate + CoA 13. Explain why ammonia is toxic. Explain the elements of inter-organ metabolism that contribute to the self-perpetuating nature of ammonia intoxication. It alters the production of neurotransmitters in the brain and reduces intracellular ATP. It causes swelling of the brain to the point that it squeezes out the bottom of the brain (liver failure). (Remember, it is very membrane permeable; it can cross the blood:brain barrier.) Elevated NH4+ concentration due to liver failure stimulates glucagon release in the pancreas, which stimulates gluconeogenesis from amino acids in the kidney, resulting in more NH4+. This ammonium stimulates more glucagon release, which stimulates release of more glucose (gluconeogenesis), and glucose promotes insulin release from the pancreas. This insulin stimulates uptake of branched chain amino acids by muscle, where they are used as oxidizable substrates. Their metabolism releases even more NH4+. This elevated NH4+ concentration in the brain stimulates glutamine synthesis, which in turn promotes tryptophan transport, which is already increased through the transporter for neutral amino acids because their serum levels decline as they are metabolized in muscle. Elevated tryptophan increases the synthesis of 5-OH-tryptamine, which is a reactive oxygen species adding oxidative stress in the brain. 14. What TCA cycle intermediate is depleted in the brain during ammonia intoxication and how does this happen? What amino acids are depleted in the brain? What are the consequences of depletion of these metabolites for the brain? α-ketoglutarate is depleted in the brain. Glutamate is also depleted. There is not as much α-ketoglutarate the flow through the TCA cycle for ATP production, there is not as much glutamate for the synthesis of γ-amino butyric acid (GABA), ATP utilization is increased, and there is increased oxidative stress due to the presence of reactive oxidative species. 15. How is ammonia intoxication treated and how do these treatments work? Diet – low protein, high carbohydrate diet reduces NH4+ production Levulose – bacteria in the colon metabolize levulose to acidic products, which promotes ammonia excretion in the feces as NH4+, which is less readily absorbed than NH3 in the gut. Antibiotics – to kill intestinal ammonia-producing bacteria Sodium benzoate and sodium phenylbutyrate – form covalent products with glycine (hippurate) and glutamine (phenylacetylglutamine). These compounds promote nitrogen excretion in the feces. 16. Explain the distinction between a ketogenic vs. a gluconeogenic fate of an amino acid. Which amino acids are exclusively ketogenic and to what are they ultimately metabolized? Ketogenic amino acids yield precursors to fatty acids or ketone bodies. They are metabolized to acetyl-CoA and acetoacetyl-CoA. Leucine and lysine are exclusively ketogenic amino acids. Glucogenic amino acids yield precursors that can be used of gluconeogenesis. They are metabolized to pyruvate, α-ketoglutarate, succinyl-CoA, fumarate, or oxaloacetate. 17. To what TCA cycle intermediates are individual amino acids metabolized and where do these metabolites enter the TCA cycle? 18. What metabolites accumulate in individuals with a defect in the branched chain keto acid dehydrogenase? What amino acids are metabolized by the pathway involving this enzyme? Α-ketoisocaproate, α-keto-β-methylvalerate, and α-ketoisovalerate. Leucine, isoleucine, and valine are metabolized by this pathway. Causes severe mental retardation. It’s a rare, recessive mutation isolated in Asian and Caucasian (Mennonites) communities. 19. What metabolites accumulate in alkaptonuria and what enzyme is defective in this disease? How are the clinical manifestations of this disorder related to the accumulation of this metabolite? Homogentisate oxidase is defective, causing homogentisate to accumulate. Patients have a damage to joints, calcifications in the urinary tract and cardiovascular system, and a reddish tint (ochre) to the skin. Urine also turns black upon standing at room temperature for 24 hours. All of this is due to accumulation of homogentisate. 20. How is S-adenosyl methionine synthesized? Methionine and ATP are added via methionine adenosyl-transferase to produce sadenosylmethionine (SAM). 21. How is methionine regenerated from homocysteine? What coenzyme is involved in this reaction? What other enzyme catalyzed reaction in mammals utilizes this coenzyme? N5-CH3 THF is converted to THF via methionine synthase. B12 is required for this reaction. The interconversion of methylmalonyl-CoA and succinyl-CoA utilizes methylmalonyl-CoA mutase, which also requires coenzyme B12. B12 dependent enzymes catalyze intramolecular or intermolecular group transfer. 22. What enzyme is defective in individuals with homocystinuria? What metabolite accumulates? How is this disease usually treated? Cysathionine β-synthase, which is a vitamin B6 dependent enzyme. Homocysteine accumulates. Two kinds of treatment: vitamin responsive and unresponsive. Vitamin responsive can be treated with pyridoxine (vitamin B6), folic acid, and vitamin B12. Vitamin unresponsive is treated with a methionine restricted diet supplemented with cystine and betaine (promotes remethylation of homocysteine to methionine). 23. What is the nature of the defect in cystinosis and how is this disorder treated clinically? There is a defect in a cystine transporter (cystinosin) in the lysosomes, so cystine accumulates there. There is often a deposition of cystine crystals in tissues (like kidneys and eyes). Treated with cysteamine, which reacts with cystine in the lysosomes to form cysteine-cysteamine; this can then be transported out of the lysosomes through a transporter for lysine. 24. How are homocysteine thiolactone and S-nitrosohomocysteine formed? Homocysteine is formed from methionine through S-adenosylmethionine. Homocysteine thiolactone is made from homocysteine by methionyl-tRNA synthetase. S-nitrosohomocysteine is formed from homocysteine and NO. Both of these react with proteins and are cytotoxic. (They react with protein amino groups, rendering the modified protein immunogenic.) 25. Which amino acids can be made from intermediates of a central pathway and what intermediate gives rise to which amino acids? From intermediates of a central pathway: Phosphoglycerate → Serine Phosphoglycerate → Serine → Glycine Citrulline → Argininosuccinate → Arginine (urea cycle) From other amino acids: Phenylalanine → Tyrosine Glutamate → Proline Methionine →Cysteine 26. Explain the role of the small intestine and kidney in the synthesis of arginine. Small intestine – citrulline is synthesized in the enterocytes; involves the synthesis of ornithine by transamination of glutamate γ-semialdehyde followed by reaction with carboamoyl phosphate (beginning of urea cycle). Citrulline is then carried by the circulation to the kidney. Kidney – citrulline taken up by kidney from circulation. Citrulline is combined with aspartate and ATP to produce argininosuccinate. The kidneys do not have arginase to complete the urea acid cycle, however; they have argininosuccinate lyase, which produces fumarate and arginine. This whole pathway is called the intestinal-renal axis. 27. What reaction is catalyzed by phenylalanine hydroxylase and what is the role of tetrahydrobiopterin in this reaction. What is phenylketonuria? Phenylalanine → Tyrosine is catalyzed by phenylalanine hydroxylase. Tetrahydrobiopterin is the required coenzyme and acts as the immediate reductant that gives up its H. It is then regenerated by NADH + H+. Phenylketonuria is when phenylaline hydroxylase is defective, or occasionally the dihydrobiopterin cofactor. If left untreated it can result in mental retardation. Affected individuals are placed on a phenylalanine restricted, tyrosine supplemented diet; this must be maintained until development is complete and while pregnant. 28. How is nitric oxide made? What are some of the functions of this short-lived molecule? Arginine + O2 via nitric oxide synthase produces citrulline. There are 3 isozymes, each with different functions: iNOS or NOSI – inducible form – immune and inflammatory responses nNOS or NOSII – neurotransmitter in GI tract, in penile erection, sphincter relaxation eNOS or NOSIII – endothelial cells – blood flow, blood pressure platelet activation EXTRAS: METABOLISM OF AMINO ACIDS Amino acids metabolized to pyruvate: Glycine cleavage system – reversible step in vitro, but in vivo it functions primarily in serine and glycine catabolism and N5N10CH2THF production. Serine – major route of serine catabolism and the principal source of glycine: – relatively minor route of serine catabolism: Threonine: Valine, Isoleucine, and Leucine – all are changed to an α-keto-ate via a transaminase and then lose CO2 and add CoASH via a branched chain keto acid dehydrogenase complex. (Don’t worry about structures or intermediates.) Lysine – exclusively ketogenic. Forms sarccharopine using α-ketoglutarate, then gives rise partly to 2-amino-adipate, which is converted to acetoacetyl-CoA. Phenylalanine and Tyrosine – both are ketogenic Cysteine: SYNTHESIS OF AMINO ACIDS Asparagine – Glutamine is the donor in animal cells, NH3 is the N donor in bacteria and plants: Glutamine: Arginine – covered in the objectives. (Split between the small intestine and kidneys.) Tyrosine – covered in the objectives. (The role of phenylalanine hydroxylase.) Cysteine: Proline – from glutamate Serine – derived from a glycolytic intermediate, 3-phosphoglycerate Glycine – source of most of the glycine required for other biosynthetic processes, including protein synthesis and pyrimidine synthesis. Also contributes to the one carbon pool (N5N10CH2THF)