BMB 462 Lecture 15 Outline of Last Lecture I Continuation of the Urea Cycle II Connections between the Urea Cycle and the TCA Cycle III Regulation of the Urea Cycle IV Metabolism of Carbon Skeletons from amino acids V Carbon Transfers VI Phenylalanine Degradation VII The Nitrogen Cycle VIII Nitrogen Fixation Outline of Current Lecture I II III IV V VI Ammonia incorporation into Carbon Compounds Amino Group Transfers Amino Acid Anabolism Important biomolecules derived from Amino acids Hormonal Signaling Hormone Classification Current Lecture Concepts to remembers from previous courses lectures I Ammonia incorporation into Carbon Compounds a The system that moves Nitrogen around is called GS GOGAT system It s the major route for incorporating Nitrogen i This is happening in plants Humans can t incorporate free Nitrogen 1 We get it already synthesized in Amino Acids from the protein in our diet 2 Plants and bacteria need to regulate the incorporation of Nitrogen ii Need glutamate to donate Nitrogen for aminotransferase b Glutamine Synthetase These notes represent a detailed interpretation of the professor s lecture GradeBuddy is best used as a supplement to your own notes not as a substitute i Glutamate NH4 ATP Glutamine ADP Pi H ii ATP is the energy source Glutamine is the nitrogen donor nitrogen is then used for synthesis of other molecules 1 Enzyme that does this is amidotransferase iii Removing the NH3 returns glutamine to glutamate so you can restart cycle and glutamine synthetase can pick up free ammonia and drop it off with transferase iv Aminotransferase uses glutamate as substrate c Glutamate Synthase i Glutamine alpha ketoglutarate NADPH H 2 glutamates NADP ii Glutamine from above becomes substrate in synthase reaction iii Transfers gamma C from side chain to alpha ketoglutarate which then provides two glutamates 1 Requires reducing power so NADPH is source of e2 Now have 2 glutamates to use in aminotransferase reactions One is source for regenerating glutamine a The other one goes down to aminotransferase and donates an amino group for synthesis When you drop it off regenerates alphaketoglutarate acceptor d Glutamate Dehydrogenase i alpha ketoglutarate NH4 NADPH H Glutamate NADP H2O ii Uses alphaketoglutarate and free ammonia reducing power to get glutamate which can then serve as glutamate used in generating glutamine iii Humans use glutamate dehydrogenase to run towards alphaketoglutarate and release free ammonia in the liver for urea cycle In plants the process can run to generate glutamate to incorporate free Nitrogen 1 This is a fairly minor route in the GDH GS system e Regulation of Ammonia Incorporation regulation in E coli i 3 levels of regulation 1 Allosteric based on quick needs in the cell regulated by levels of products 2 Controlling enzymes already there by covalent modification 3 Producing more enzymes ii 2 parts to allosteric regulation in bacteria 1 Levels of glycine and alanine tell cell whether you have enough Amino Acids When levels start increasing there s going to be allosteric inhibition of glutamine synthetase 2 Inhibition of the acceptors for amidotransferase Glutamine is the e donor As glutamine increases the levels of synthetase reduce iii The cell is incorporating just enough nitrogen to fulfill functions without wasting Energy on gathering excess iv Covalent Regulation of Glutamine synthetase 1 Adenylylation Adding AMP to enzyme analogous to phosphorylation and dephosphorylation 2 Uridylylation is adding UMP a Both used to control various proteins 3 An indicator that there is enough Nitrogen is plenty of glutamine and lots of phosphate a Means you broke down a lot of ATP so don t have much energy left b Don t want to be undergoing biosynthesis so glutamine synthesis gets an AMP added and becomes inactive 4 Layering this adenylylation with allosteric control is very sensitive regulation v Transcriptional Regulation 1 When PII has UMP attached activating glutamine synthetase it also activates transcription of the GS gene a So the cell makes more of the protein as well More protein is more activity II Amino Group Transfers a Aminotransferases use the alpha amino group from glutamate and move it to the alpha keto acid b Amidotransferases use side chain gamma amino group from glutamine transfer it to activated hydroxyl or a keto group i Donate Nitrogen to one or the other acceptor so that the product has Nitrogen incorporated III Amino Acid Anabolism a Synthesis is Ketone groups to Amino Acids Breakdown is Amino Acids to ketone groups b Anabolism is not simply the reverse of catabolism The carbon is not coming from the same place it was broken down to i Some reactions in common but also have unique steps c Diverging pathways start with a few intermediates 7 of them and from them you make all 20 Amino Acids so there has to be branches in pathway to make all of the Amino Acids d General characteristics i Take carbon from central metabolism ii Modify the carbon skeletons to get alpha keto acids 1 This uses a lot of ATP for activation in order to do biosynthesis 2 Perform reduction of carbon which will frequently require NADPH as e donor Many reactions also require PLP this allows us to do chemistry around the alpha beta and gamma carbons of amino acids iii After you get alpha keto acid add an amino group using an aminotransferase Use glutamate as a donor and move it to a keto group in alpha keto acid iv Additional modifications are then performed to get to the correct final amino acid product e Sources of Carbon Skeletons i The core sources of carbon are from pentose phosphate pathway 1 The ribose and erythrose phosphates ii Glycolysis 3phosphoglycerate phosphoenolpyruvate and pyruvate iii TCA alpha ketoglutarate and oxaloacetate 1 Only these 2 whereas breakdown had almost all intermediates going to TCA Cycle f Transamination aminotransferases add N g Biosynthetic Families i Know precursors alpha ketoglutarate and oxaloacetate from TCA pyruvate 3PG PEP from glycolysis pentose phosphates ii Keto amino acid pairs Alphaketoglutarate and glutamate oxaloacetate and aspartate pyruvate and alanine h Essential vs Non essential amino acids i Non essential the body can make enough of the amino acid that you don t need to include it in the diet ii Essential The body can t make the amino acid because it doesn t have the enzymes needed or can make it but not in the quantities needed by the body 1 They must be included in the diet 2 i e arginine in urea cycle i
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