BIOCHEMISTRY
Module No.
Biosynthesis of Nutritionally NE AA
8.1.1
Dr. Pacifico Eric E. Calderon, M.D.
February 15, 2017 *Except when replacing protein losses, nitrogen equilibrium can equilibrium can be maintained at any level of protein intake ABOVE requirements. A high intake of protein DOES NOT lead NOT lead to positive nitrogen balance; although it increases the rate of protein synthesis, synthesis, it also increases the rate of protein catabolism,, so that nitrogen equilibrium is maintained catabolism albeit with a higher rate of protein turnover. * Both protein synthesis and catabolism are catabolism are ATP expensive, and this increased rate of protein turnover explains the increased diet-induced thermogenesis seen in people consuming a high protein diet.
Outline
I. Nitrogen Metabolism II. Amino Acid Acid Catabolism III. Excretion of Excess Nitrogen IV. Removal of Nitrogen LEGEND Lecture Powerpoint, Audio Powerpoint, Audio /Transers’ notes, notes, Textbook I. Nitrogen Metabolism Nitrogen Balance
A state where the amount of nitrogen ingested each day is balanced by the amount excreted, resulting in no net change in the amount of body nitrogen Nitrogen intake should equal nitrogen excretion Nitrogen Balance = Intake – Excretion The state of protein nutrition can be determined by measuring the dietary intake and output of nitrogenous compounds from the body. Although nucleic acids also contain nitrogen, protein is the major dietary source of nitrogen nitrogen and measurement of total nitrogen intake gives a good estimate of protein intake. Nitrogen from the body is mainly in urea & Nitrogen from urea & smaller quantities of other compounds in urine, undigested protein in feces, feces, significant amounts may also be lost in sweat & shed skin.
3. Protein Turnover
4. Amino Acid Pool
1. Positive Nitrogen Balance
Intake > Excretion Net accumulation of proteins as in growth and pregnancy, and as in person in recovery from protein loss In other words if your intake is greater it follows that
excrete them because you use them up to make proteins.
Intake < Excretion Net breakdown breakdown of protein protein as in surgery, advanced cancer, kwashiorkor or marasmus, starvation, diet deficient in essential AA (promotes CHON degradation), degradation), aging, trauma, infection So, if your intake is lesser and the rate of excretion remains unaffected it would also mean that mas madami ka naExcrete kesa sa nakain mo. So saan nanggaling yung extra na naExcrete if kulang na nga yung intake mo? Sa degradation of the proteins in your body like sa muscle and you may also find this in hypermetabolic states.
Transers: CARLOS, HARINA, HUEVAS, GAYADOS
Sum of Sum of all free amino acids in cells and ECF Three possible sources: Degradation and turnover of body o protein (endogenous), most of which are reutilized Dietary intake (exogenous) o Synthesis of Synthesis of nonessential amino acids o There is no storage form for amino acids, Thus everything in excess is degraded
S ynthes i s > deg radation . When you eat, you do not
2. Negative Nitrogen Balance
Refers to CHON synthesis and degradation 300-400 g per day Amount of protein degraded and resynthesized from amino acid (AA) While approximately 75% of the AA liberated AA liberated by protein degradation degradation are utilized utilized,, the excess free amino acids are not stored for stored for future use. AA NOT NOT immediately immediately incorporated incorporated into new protein are protein are rapidly degraded The major portion of the carbon skeletons of AAs is converted to amphibolic intermediates, amphibolic intermediates, while in humans the amino nitrogen nitrogen is is converted to urea urea and and excreted in the urine.
Remember that proteins have no storage form , so behind ind hig h protein protein diets protei protei n this is the logic beh
comprise the calories in the diet, but are degraded eventually * Conversely, the amino pool is depleted by three routes: 1. synthesis synthesis of of body protein 2. consumption consumption of of amino acids as precursors of precursors of essential nitrogen-containing small molecules 3. conversion conversion of of amino acids to glucose, glycogen, fatty acids, and ketone bodies, or oxidation of CO2 + H2O *Although the amino acid pool i s small (comprising about 90-100g of amino acids) in comparison with the amount of protein in the body (about 12kg in a 70kg man), it is conceptually at the center of whole-body nitrogen metabolism. Page 1 of 6
BIOCHEMISTRY
PROTEIN METABOLISM
Module 8, Lecture 1.1
In healthy, well-fed individuals, the input to the amino acid pool is balanced by the output. That is, if the amount of AAs contained in the pool is constant. The AA pool is said to be in a ready state, and the individual is said to be in nitrogen balance.
Digestion of Proteins
SUMMARY
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1.
Digestion begins in the stomach with HCl (from parietal cells) and pepsin (from chief cells) Acid activates pepsinogen to pepsin Pepsin capable of auto-activating itself
Picture above: Just shows the summarized Synthesis and degradation of Proteins. Take note of the amount of proteins.
MAR K ING :
Ubiquitin-proteosome
mechanism
(energy dependent) binds to endogenous protein (proteins that are created intracellularly) that needs to be degraded by proteos ome pathway (GARBAGE disposal) energy dependent manner – alpha carboxyl of glycine of ubiquitin to lysine amino group of protein substrate Exogenous (extracellular) degraded in the lysosomes (nonenergy dependent manner)
HCL functions to kill bacteria and aid in denaturation of proteins making it more susceptible to hydrolysis by some proteases Pepsin chief cells or serous cells (pepsin is endopeptidase, acid stable) secreted as an inactive zymog en / proenz yme ; activated by cleavage of extra AA ZYMOGENS: Inactive form of enzymes o
2.
Digestion by pancreatic enzymes that are i nitially secreted as zymogens. Trypsin is the common activator. Trypsinogen Trypsin Via enteropeptidase Endopeptidases Trypsin Chymotrypsin Elastase
Exopeptidase Carboxypeptidase Aminopeptidase
Elastase, carboxy, chymo cleavage sites.
all
of these have specific
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Aminopeptidases in the brush border liberate amino acids and dipeptides
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Free amino acids are absorbed by secondary active transport
Secondary active transport like glucose and galactose. Fructose is via facili tated diffus ion.
What are the final products that get absorbed in the small intestinal mucosa? BOTH free amino acids and di-/tripeptides.
Pi cture above: Shows what contributes to the AA pool
Transers: Zia, Dara, Sam, Billy
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BIOCHEMISTRY
PROTEIN METABOLISM
II. Amino Acid Catabolism
The presence of an alpha-amino group keeps amino acids safely locked away from oxidative breakdown
Module 8, Lecture 1.1
Uricotelic
IV. Removal of Nitrogen
Seen in birds, which excrete uric acid as semisolid guano
2 main steps in removing nitrogen from AA o Transamination o Oxidative Deamination
Alpha-amino group functions to keep the AA acid safe thus removing the alpha amino group is an OBLIGATORY step in the catabolis m of all AA The biggest step that must be undertaken if you want to use your amino acid is to remove the alpha amino g roup. There are two ways either you put it out/take it away or you transfer it to another.
First Phase of AA Catabolism
Removal of the α-amino group (a process called deamination) forming : ammonia and a corresponding α-ketoacid (whole structure pointed) nahawa ng histo. haha!
Sabi kasi walang forever eh….maghihiwalay din kayo. <3
What happens to ammonia (NH3)? Maybe excreted as free ammonia in urine and stool Majority is converted to urea before being excreted in urine (urea is the major disposal form of nitrogen) Second Phase of AA Catabolism Carbon skeletons of α-ketoacids are converted to common intermediates of energy-producing metabolic pathways o Glycolysis o Krebs Cycle
Step 1: Transamination The funneling of amino groups to glutamate Occurs in all cells of body All amino acids must transfer their amino groups α-ketoglutarate to to form glutamate (Exceptions: lysine and threonine DEAMINATION) Enzymes: Aminotransferases (formerly transaminases) Found in the cytosol of cells throughout the body (liver, kidney, intestine) Alanine aminotransferase (ALT) Aspartate aminotransferase (AST)
Co-enzyme: Pyridoxal Phosphate (Vitamin B6)
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1 phase: throw away amino group sa urea cycle nd 2 phase: recycle whats left of AA which is the alpha keto acid which will become the carbon skeleton
III. Excretion of Excess Nitrogen Urea is relatively move toxic than uric acid, that’s why is has to be diluted in urine in the body. Uric acid is not very toxic, and may be concentrated to a semi-solid paste without causing toxic effects.
Ammonotelic Seen in telostean fish, which excrete highly toxic ammonia Ureotelic
4 So initially we have an (1) AA from protein catabolism and an (2) alpha ketoglutarate from the Krebs cycle. So when this two interact together with the help of the (3) aminotransferase the amino grp. of the AA which is the ammonia(NH3) will be transferred to alpha ketoglutarate forming now our (4) g lutamate. This process is vice versa, meaning we can also form AA especially NE AA or alpha ketoglutarate if glutamate & alpha keto-acid interacts together.
Transamination process is true for all AA EXCEPT for LYSINE & THREONINE because they lose their amino group via DEAMINATION and NO T by Transamination.
Seen in land animals, including humans, who excrete non-toxic, water-soluble urea
Transers: ZIA, DARA, SAM, BILLY
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BIOCHEMISTRY
PROTEIN METABOLISM
Module 8, Lecture 1.1
Enzyme: glutamate dehydrogenase Provides α-Keto acids that can enter the central pathways of energy metabolism and ammonia
Transamination only exchanges amino groups. NO FREE AMMONIA
Remember the pairings: Glutamate Alanine Aspartate
= = =
α-ketoglutarate pyruvate oxaloacetate
Almost all amino acids transaminate with glutamate and α-ketoglutarate
Oxidative deamination actually LIBERATES AMMONIA
* α-Ketoglutarate plays a pivotal role in AA metabolism by
accepting the amino groups from most AAs, thereby becoming GLUTAMATE. *Glutamate produced by transamination can be oxidatively deaminated or used as an amino group donor in the synthesis of nonessential AAs. This transfer of amino groups from one carbon skeleton to another is catalyzed by a family of enzymes called aminotransferases (aka transaminases) These enzymes are found in the cytosol and mitochondria of cells throughout the body * All AAs, except lysine and threonine, participate in transamination at some point in their catabolism.
Clinical Correlate ALT is also known SGPT (serum glutamate: pyruvate transferase) Pyruvate and alanine interconvert with transamination
Pyruvate (alanine becomes pyruvate pag natanggalan ng amino group) glutamate-pyruvate transaminase
Ammonia enters the UREA CYCLE to become urea (less toxic more soluble form of From the picture above: Glutamate further undergoes oxidative deamination para completely matangal ang nitrogen group as ammonia na. wala ng sasalo sa kanya. This now enters the urea cycle. This transamination – oxidative deamination happens in your body. Ammonia is toxic. Has to be transported to the area where urea cycle occurs so it can be converted to urea.
Excess nitrogen from the peripheral tissues can also reach the liver through glutamate
AST is also known as SGOT (serum glutamate: OAA transferase) Aspartate and oxaloacetate interconvert with transamination
oxaloacetate (aspartate becomes OAA) glutamateoAA transaminase
AST & ALT are commonly used to test for liver function. So it is used as a maker to indicate if the patient has a liver disease which is indicated by an increased in AST & ALT (more specific for liver than AST)
Glutamate + Ammonia Glutamine Enzyme: Glutamine Synthetase Glutamine serves as the NONTOXIC transport form of ammonia
Step 2: Oxidative Deamination Occurs in the liver and kidney only Only for glutamate Glutamate is oxidized and deaminated to yield free ammonia (NH3) which is used to make urea
Transers: Zia, Dara, Sam, Billy
So ang g lutamine ay glutamate na nadagdagan lang ng isa pang nitrogen group.
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BIOCHEMISTRY
PROTEIN METABOLISM
If you think about it, glutamine is the non toxic form of ammonia for most tissues. Glutaminase – breakdown at liver to release ammonia Alanine: transaminated pyruvate (occuring only in muscles)
What about the muscle?
Excess nitrogen from the peripheral tissues can also reach the liver through alanine, especially in muscle
Module 8, Lecture 1.1
Mus cle i s not capable of g luconeog enes is – ONLY the liver can do this. Pyruvate (which is a substrate for gluconeogenesis) is converted to alanine first. Alanine shuttles pyruvate to the liver where gluconeogenesis can take place.
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ammonia removed from alanine then enters the urea cycle. pyruvate is used by the liver for gluconeogenesis produced glucose is then exported to other tissues
Glutaminase Pyruvate + Glutamate Alanine + α- ketoglutarate Enzyme: alanine aminotransferase (ALT or SGPT) Yung pyruvate naman, dagdagan mo lang ng amino group, alanine na.
Deaminates glutamine to produce ammonium ion + (NH4 ), which is excreted from the body Present in two tissues: kidney and small intestines
KIDNEYS SMALL INTESTINES Eliminates ammonium ion in Ammonium ion sent to the urine liver via the portal circulation for the urea cycle Kanina diba glutamate to glutamine happens in tissues, muscle. E pano naman pag tatanggalin na ang nitrogen sa glutamine. Kasi itatapon na siya from the body. Eto na yun. So reverse lang eto nung kanina. Ang difference lang, eto nangayayari sa kidneys and the intestines.
In the muscle, there is transamination of pyruvate – end product of aerobic glycoysis (remaining alpha keto acid is alanine). Once alanine reaches the liver, it is transminated again to form pyruvate; glutamate will undergo oxiative deamination via glutamate dehydrogenase to liberate free ammonia to enter urea cycle
Where is Glutamine synthase found? Liver, Muscle and CNS
Pyruvate can enter gluconeogenesis to produce glucose
Where is GLUTAMINASE found? Kidneys and Small Intestine
Transers: ZIA, DARA, SAM, BILLY
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BIOCHEMISTRY
PROTEIN METABOLISM
Module 8, Lecture 1.1
IN OTHER WORDS Glutamine Synthethase is for transporting ammonia Glutaminase is for getting rid of ammonium ion via the Urea Cycle SUMMARY Amino Group/ NH3
Reference REVIEW
Dr. Calderon’s Powerpoint Harper ’s Illustrated Biochemistry 30th Edition th Lippincott’s Biochemistry 6 Edition
Q: What is another substance, important in vitamin B12 metabolism that is also produced by parietal cells? A: Intrinsic Factor Q: Can you name other substances absorbed by secondary active transport in the small intestine? A: glucose and galactose Q: After absorbing the products of protein digestion, which of the following products can you see inside enterocytes? A. B. C. D.
Amino Acids Dipeptides Tripeptides All of the above
Q: While all amino acids have nitrogen groups in their structure, not all amino acids can transport nitrogen groups to the liver to enter the urea cycle. Which of the following amino acids cannot transport amino acids to the liver? A. B. C. D.
aspartate glutamate glutamine alanine
Q: What do you call the metabolic pathway whereby lactate produced during anaerobic respiration in muscles is reconverted to glucose in the liver? a. b. c. d.
Transers: Zia, Dara, Sam, Billy
Embden-Meyerhoff Pathway Bohr Effect Cori Cycle Tricarboxylic Acid Cycle
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