Exam IV

• Sources:
• dietary protein
• body protein (muscle)
• Digestive juices

• Uses:
• conversion to purines, pyrimidines, heme, creatine, amines
• degredation for gluconeogenesis, energy, storage as glycogen or fat
• protein synthesis
• excretion in feces
• deposits in skin

• -food enters the stomach and gastric juice is secreted fromt he glands int he stomach
• -gastric juice includes pepsinogen and hydrochloric acid
• -pepsinogen is secreted with a prosegment bound to the active site
• -the low pH causes the prosegment to come off and pepsinogen is activated to become pepsin
• -pepsin breaks down the proteins into large polypeptides
• -the chyme enters the duodenum and secretin and cholecystokinin are released from specialized endocrine cells into circulation
• -gall bladder releases alkaline secretion
• -pancrease secretes trypsin, chymotrypsin and elastase which break things up into di and tripeptides
• -enzymes on or within the intestinal epithelial cells degrade them down into amino acids
• -then the proteolytic enzymes are degraded

• -digestive enzymes are secreted as inactive zymogens in the small intestine
• -cholecystokinin is a hormone that causes enteropeptidase to be secreted from the luminal surface of th SI
• -enteropeptidase clips a hexapeptide from the N-terminus of trypsinogen which activates it to trypsin by forming the catalytic triad
• -trypsin plays a central role in protein digestion regualtion since it activates other pancreatic zymogens including more trypsinogen

• inflammation of the pancreas caused by active trypsin
• characterized by pain in the upper middle and upper left part of the abdomen
• Herditary: a mutation in the amino acid sequence of the trypsin molecule itself so that the inhibitor cannot bind to the active site to shut it off while still inside the pancreas
• Non-hereditary: blockage of the pancreatic duct cause trypsin to accumulate in the pancreas and htere is not enought inhibitor to keep it all inactive

• usually taken up against concentration gradient using active transport
• Na+-coupled transporters are specific for groups of amino acids rather than for individual ones
• the Na+ gradient is maintined by the Na/K ATPase; so the transporters can move amino acids into the cell while utilizing the energy released from moving Na+ into the cell with it
• a facilitated transporter on the serosal (basal) side passes the amino acid into the blood stream

• cysteine in the urine
• caused by a defective membrane transporter in the kidney glomeruli
• the cysteine is readily oxidized tot he less-soluble form of cystine which can cause painful kidenys tones

• a defect in an amino acid transporter in epithelial cells that results in the excretion and loss of several neutral amino acids in the urine
• also deficient in absorption in the intestine
• results in a tryptophan deficiency which is needed to make neurotransmitters and niacin for NAD and NADP
• leads to Pellagra, characterized by the three D's: dermatitis, dementia, and diarrhea

• 1. Ammonia: only a very small amount in the blood and urine
• 2. Urea: 90% of excess nitrogen is excreted as urea; excretion is also a direct reflection of protein intake (in nitrogen balance of a healthy adult)
• 3. Uric acid: only a little in the urine as a result of purine breakdown
• 4. Creatinine: small amounts in urine; proportional to muscle mass

• Positive: the body retains more of the protein nitrogen than it excretes as urinary nitrogen; happens when body protein is accumulating such as during childhood or pregnancy
• Negative: more nitrogen is excreted than is ingested; during starvation or inadequate protein nutrition; associated with fever, speticemia and trauma; high quality protein meals are necessary for patients recovering from these conditions

• Step 1: transamination shuffle; catalytic transfer of the alpha-amino group of an amino acid to a carrier alpha-keto acid by transaminase; all transaminases require pyridoxal phosphate (vitamin B6) as a coenzyme; major alpha-keto acids used are alpha-ketoglutarate, pyruvate, and oxaloacetate (converted to glutamate, alanin, and apsartate, respectively); this reaction is reversible so it can be used to make many amino acids
• Step 2: the glutamate dehydrogenase reaction; release of the transferred alpha-amino nitrogen from the carrier as ammonia; the enzyme is found only in the mitochondria; converts NAD to NADH; if this is in the liver, the ammonia can be converted to urea; reversible and controlled by ATP/ADP ratio (more ATP, slow rxn)
• Step 3: Ammonia Fixation; via the reverse of the glutamate dehydrogenase reaction depending on tissues needs; via glutamate synthetase reaction which converts glutamate to glutamine using ATP and ammonia (irreversible); this second reaction is important because glutamine can be used to make asparagine, purines, and pyrimidines and can also be used as fuel for the intestines and kidneys; this reaction is also important for regulating blood pH in the kidneys

in the liver and to a small degree in enterocytes

• Mitochondria: creation of carbamoyl phosphate by carbomoyl phosphate synthetase (CSPI), this is the regulated step via availability of NAG; ornithine + carbomoyl phosphate --> citrulline
• Cytosol: citrulline + aspartate --> arginosuccinate--> fumarate (released) + arginine--> urea (released) + ornithine

Ordinarily Careless Crappers Are Also Frivolous About Urination

• N-Acetyl Glutamate
• an intermediate in the synthesis of carbamoyl phosphate
• only found in the mitochondria
• synthesized by acetyl-CoA and glutamate by NAG synthase which is stimulated by arginine, a downstream product (example of a feed-forward mechanism)

• clinical features: developmental delays, mental retardation, seizure, protein intolerance
• diagnosis: blood/urine test for ammonia; high arginine levels indicate arginase deficiency
• treatment: restrict protein intake; supplement with arginine (an essential aa); try to increase nitrogen excretion

• 1. aspartate/oxaloacetate
• 2. glutamate/alpha-ketoglutarate
• 3. fumarate/malate

• After breakdown, the carbon skeletons of glycogenic aa's are intermedaites of glycolyusis or the TCA cycle and can contribute to the synthesis of glucose and glycogen via gluconeogenesis or glyogenesis
• the breakdown products of ketogenic aa's give rise to short chain fatty acids that undergo beta-oxidation to form ketone bodies and acetyl-CoA
• some amino acids can be both
• NOTE: leucine and lysine are the only two aa's that are stricly ketogenic; some amino acids have multiple entry points, ex. threonine and tyrosine

• Alanine, serine, and cystein
• glycine via serine
• tryptophan via alanine
• threonine via glycine via serine
• these reactions are dehydrations followed by deaminations

• glutamate
• glutamine, arginie and proline via glutamate

• threonine, methionine, isoleucine, and valine
• all via an alpha-ketoacids
• used amino transferases in the first step (of some) and dehydrogenase in the second step to go from the metoacid to propionyl-CoA, which becomes succinyl-CoA, which becomes Succinate
• the reaction requires lipoate, thiamine and FAD

• caused by a deficiency in the branched chain 2-ketoacid dehydrogenase that converts the ketoacid intermediates of amino acid breakdown into propionyl-CoA to become Succinate
• isoleucine, leucine, valine, and alpha-ketobutyrate accumulate and make the urine smell like maple syrup
• commonly observed with mental retardation
• treatment: severly restrict the intake of branched amino acids

• via interconversions between the D and L forms of methylmalonyl-CoA
• one of the enzymes, methylmalonyl-CoA mutase, needs the cofactor adenosylcobalamin which is dervied from vitamin B12
• defects in any of the three enzymes involved in this reaction have been implicated in some forms of ketoacidosis; many are caused by a deficiency in vitamin B12 or the inability to convert it to the cofactor needed
• may see methylmalonate in the urine, especially if there is a defect in the intrinsic factor needed toabsord B12 in the stomach

• aspartate
• asparagine via aspartate
• note: the ammonia from aspartate is transferred to an alpha-ketoglutarate to make glutamate

• phenylalanine is converted to tyrosine which is converted to fumarate and acetoacetate
• note: both of these are essential amino acids
• note: the products are glucogenic and ketogenic

• Phenylketonuria, a hereditary defect in the metabolism of phenylalanine
• usually associated with a mutated form of phenylalanine hydroxylase which converts Phe to tyrosine in the liver or the inability to make tetrahydrobiopterin (BH4) which is needed for the conversion
• clinical features: irreversible mental retardation, tremors, hyperactivity, light pigmentation in skin and eyes
• phenylalanine accumulates and enters other pathways to make products that cause the urine to smell musty
• treatment: dietary restriction of phenylalanine supplemented with tyrosine

• deficiency in homogentisic acid oxidase
• homogentisic acids is an intermediate between tyrosine and fumarate/acetoacetate
• characterized by dark urine and arthiritis

• Step 1: glucose-6-phosphate becomes Ribose-5-phophate via the pentose-phosphate shunt
• Step 2: ribose-5-phosphate becomes PRPP via PRPP synthetase and ATP <-- comitted/regulated step
• Step 3: PRPP picks up an amine from glutamine to become 5-phosphoribosylamine using glutamine phosphoribosyl pyrophosphate amidotransferase<-- inhibited by the end products
• Step 4: 5-phosphoribosylamine becomes inosine (hypoxanthine with ribose-P) aka IMP; uses amino acids as amine donors, and folate and biotin for carbons; the process includes ring closure and needs ATP
• Step 5: IMP+ATP=GMP or IMP+GTP=AMP; this system allows for equal pools of each to be formed
• Step 6: conversion of hte monophosphates to diphosphates by AMP kinase or GMP kinase
• Step 7: diphosphate converted to triphosphates by nucleoside diphosphate kinase (this is also used to activate AZT)

• HGPRT pathways:
• -Guanine + PRPP = GMP
• -hypoxanthine +PRPP = IMP

• APRT pathway:
• -adenine + PRPP = AMP

• Step 1: nucelotides must lose phosphate to become nucleosides; hydrolyzed by phosphatases
• Step 2: Adenosine is deaminated by Adenosine deaminase (ADA) to make Inosine
• Step 3: Inosine broken down into hypoxanthine and ribose-1-phosphate by purine nuceloside phosphorylase (this is irreversible and does not work on deoxyadenosine)
• Step 4: hypoxanthine converted to xanthine and xanthine to uric acid by xanthine oxidase; guanine converted to xanthine by guanine deaminase and xanthine to uric acid

• GTP makes more ADP from IMP and ATP makes more GMP from IMP
• GMP and AMP inhibits the amidotransferase to block de novo synthesis
• more PRPP activates the amidotransferase to increase de novo synthesis

• high levels of uric acid in the blood
• uric acid crystals accumulate in the joints and kidneys
• Possible etiology:
• 1. partially defective HGPRT: reduced IMP and GMP from salvage so there is no negative feedback on amidotransferase so PRPP accumulates and activate de novo purine biosynthesis
• 2. PRPP synthetase less susceptible to feedbakc inhibition: no negative feedback from end products means more de novov and overproduction of PRPP
• 3. Glucose-6-phosphatase deficiency: G-6-P shuttled into the pentose-phosphate shunt; more PRPP
• Treatment: Allopurinol; analogue of hypoxanthine which is a competetive inhibitor of xanthine oxidase so less uric acid is produced

• X-linked defect in HGPRT
• clinical features: self-mutilation, mental illness, and gout due to excessive uric acid in the blood
• increase rate in de novo purine synthesis
• treatment: Allopurinol

• block folate synthesis in bacteria so they cant make purines and cant grow
• it is an analogue of a component of folic acid

• analogue of hypoxanthine used for cancer treatment
• it is salvaged by HGPRT which uses up free PRPP and slows down de novo purine biosyntehsis
• it tricks the cell into thinking it salvaged purines because the nucleotide analogue inhibits the de novo pathway

• Step 1: glutamine + ATP + HCO3 = carbamoyl phosphate (note: this is done by CPSII in the cytoplasm)
• Step 2: carbamoyl phosphate + aspartate = N-carbamoyl aspartate via acetate transcarbamoylase
• Step 3: ring closure by dihydroorotase to form dihydroorotic acid
• Step 4: oxidation by dihydroorotate dehydrogenase to form orotic acid (note: all the enzyme sup to this point are ont he same polyprotein)
• Step 5: Orotic acid + PRPP = OMP via orotate phosphoribosyltransferase
• Step 6: OMP to UMP via OMP decarboxylase
• (UMP can be converted to UTP by minases using ATP)
• Step 7: UTP + glutamine = CTP by CTP synthase

• reversible conversion of bases to nucleotides using a nucleoside phosphorylase and a kinase
• two step process: pyrimidine --> pyrimidine-ribose (nuceloside)--> pyrimidine-ribose-phosphate (nucleotide)

• nucleotides converted to nucleosides by phosphatases
• cytosine is converted to uridine by cytosine deaminase before it can proceed
• the nucelotides are converted into free bases by pyrimidine nuceloside phosphorylase
• the end products can be measured for therapeutic reasons

• All four NDPs can be reduced to form deoxyribonuceloside diphosphates by ribonucleotide reductase<--the enzyme is inhibited by the cancer drug Hydroxyurea
• electron donor is thioredoxin, which needs NADPH to regenerate it
• it is a complicated enzyme with many domains and many processes
• the process also needs FAD

• dUDP is converted back to dUMP which can become TMP via thymidylate synthase
• N5,N10-methylene tetrahydrofolate (Folate H4) is the on carbon donor and becomes Folate H2 in the reaction; dihydrofolate reductase regenerated folate H4
• Chemotherapeutic agents target thymidylate synthase and dihydrofolate reductase

De novo: CPSII (inhibited by UTP and stimulated by PRPP and ATP; important in balancing the pools); OMP-decarboxylase (inhibited by UMP, the end product; note that this is on the same protein as oratate phosphoribosyltransferase so Orotic acid builds up instead of OMP; aka Orotic aciduria)

• On the level of ribonucleotide reductase
• Overall activitof hte enzyme is stimulated by ATP and inhibited by dATP
• there is also finer adjustment of enzyme activity by the other deoxyribonucleotides to make more dATP which will eventually turn the system off
• NOTE: ADA deficiency and SCID is associated with a build up of dATP which may inhibit the synthesis of other deoxyribonucleotides which may prevent lymphocyte proliferation

• an analogue of folic acid that competatively inhibits dihydrofolate reductase
• completely inhibits TMP synthesis
• used for leukemia and other cancers

converted to fdUMP in the body and covalently binds to thymidylate synthetase to block TMP synthesis

• Dopamine: catecholamine associated with the pleasure system; tyrosine-->Dopa-->Dopamine; tetrahydrobiopterin used in the hydroxylase reaction and PLP is used the decarboxylase reaction; Parkinson's patients are treated with Dopa so they can convert it into dopamine
• Melanin: tyrosine + dopa = melanin using tyrosinase; deficiency of tyrosinase leads to albinism
• Thyroxine: two tyrosine residues on the thyroglobulin bind covalently and get iodinized

• NOTE: Folates and SAM are used on 1 carbon transfer reactions
• Serine + THF <--> Gylcine + N5, N10 methylene tetrahydrofolate; reversible reaction so it is a route for de novo synthesis of both glycine and serine; can also give rise to pyruvate
• Dopamine--> Norepinephrine + S-adenosyl methionine (SAM) --> Epinephrine + S-adenosyl homocysteine
• Glycine + arginine --> guanidinoacetic acid + SAM --> Creatine
• 5-methyl THF + homocysteine ---> THF + methionine; uses methionine synthase and methyl B12 (methylcobalamin)
• Degredation fo homcysteine: homocysteine + serine--> cystathione--> alpha-ketobutyrate and cystein; first step catalyzed by cystatione synthetase with PLP as a cofactor and the second step is catalyzed by cystationase; note that this is downstream of methionine degredation so if met is deficient, cystein is deficient
• Cysteine--> Taurine

• remember that the reaction to go from THF to 5-methyl THF if irreversible
• the only way to regenerate THF from this is via the methionine synthase reaction using homocystein and generating methionine and THF which requires B12
• So a B12 deficiency will also result in a THF deficiency and a build up of methyl-THF

• increased risk of arterial disease; long fingers; vision problems; common psychiatric disturbances
• most commonly caused by a defect in cystathionine synthase
• Treatment: reduce homocysteine while maintaining amino acid balance; with Vit B6 (PLP) supplements; supplements with choline, folate, cysteine

• In the muscle cells they are used for feul
• the liver does not have the branched chain aminotransferases but the muscle does
• ammonia is added to mak or alanine in the muscle cells which goes back tot he liver to be metabolized (Cahill Cycle)

the liver can absord glutamine from the blood for production of ammonia or it can synthesize glutamine for export as a fuel fo rht eintestine or for pH control in the kidneys via the glutaminase reaction

glycolytic and TCA cycle intermediates such as 3-phosphoglycerate and pyruvate

• Glucose-6-Phosphate used to make NADPH via pentos-ephosphate shunt that is needed to make fats
• Dihydroxyacetone needed to make the glycerol backbone
• Pyruvate becomes acetyl CoA which is used to make triglycerides, phospholipids, FA's, ketone bodies, and cholesterol

• DHAP
• Acetyl-CoA: to make ketogenic aa's, OAA, alpha-ketoglutarate for E, Q, P, D, and N

• Allosteric: very rapid response; affect the activity of enzymes that are already present and functioning (example is an increase in activity of glycogen synthase when glucose-6-phosphate is present)
• Adaptive: changes in the amount of enzyme present; much slower process but the effects last longer; due to an increase in the rate of synthesis of the enzyme or a decrease in its degredation (examples include and increase in proteases after a high protein meal)
• Covalent: adding or removing a covalently bound regulator to an enzyme; usually a phosphorylation (example is the phosphorylation of hormone-sensitive lipase under the influence of glucagon to break triglycerides down into FA's)
• Compartmental separation: different enzymes in different locations

• Insulin: promotes fuel storage after a meal and promotes growth; stimulates synthesis of glycogen, fatty acids and protein
• Glucagon: mobilizes fuels; maintains blood glucose during fasting; activates FA release from adipose tissue
• Epinephrine: mobilizes fuel during acute distress; glycogen and TAG breakdown
• Cortisol: long term regulation that promotes gluconeogenesis from proteins and mobilization of fatty acids from adipose tissue

• a chaperone protein that associates with steroid hormone receptors in the cytoplasm
• when hormone binds, hsp90 is releases and the complex translocates tot he nucleus where it binds DNA and initiates transcription

• 1. The well fed phase: exogenous fuel is supplying and maintaining blood glucose
• 2. Post-absorptive state: the overnight fast; glycogen is the main source of glucose; changes in the I/G ration have initiated gluconeogenesis from non-carb stores
• 3. Early starvation: glycogen almost depleted and proteins are beginning to be broken down for gluconeogenesis; epinephrine released which activates fatty acid release and glycogen breakdown; all tissue except liver and muscle using glucose still
• 4. Middle/intermediate starvation: decrease in total body glucose consumption and decrease gluconeogenic reactions; brain adapts and begins using ketone bodies for fuel
• 5. Late/prolonged starvation: decreased need for gluconeogenesis bc the brain is primarily running off of ketone bodies
• NOTE: nitrogen excretion in urine drops significantly; almost no urea is excreted but more nitrogen is excreted as ammonia

• glucose enters the cell via a transporter and goes through glycolysis
• increase in the ration of ATP/ADP
• ATP exits through a potassium sensitive channel so the membrane is depolarized
• Calcium can enter througha voltage dependent channel and insulin granules are released

• GLP1 and GIP; hormones produced in the intestine that go to the pancreas to stimulate beta cells to make insulin
• this is why oral glucose has a larger effect than glucose injected directly into the bloodstream
• Exenatide and Sitigliptin are analogues and act on the potassium channels to cause depolaraization and release of insulin

• 1. Genetics: IRS-1 mutation; it is part of the insulin signalling pathway
• 2. Fat: white fat secretes adiponectin which promotes insulin sensitivity, but the more fat you have the less adiponectin it makes and the less insulin repsonse
• 3. Free fatty acids: from adipose tissue, may reduce insulin sensitivity
• 4. Cytokines: adipose tissue releases TNFalpha and IL-6 which reduce sensitivity

• 1. Glucose reacts with metals to form reactive oxygen species (ROS) which can damage cells and tissues and DNA
• 2. Glucose can get converted to sorbitol which slowly gets converted to frustoce, but the second reaction is slower so sorbitol builds up and can cause dibetic retinopathy
• 3. Glycation of proteins such as hemoglobin leading to advnaced glycation products which are reactive; can be diagnostic

• Normally: it increases fatty acids synthesis in the liver; activated LPL to move FA's into cells and decreases hormone sensitive lipase to increase fat deposition in the tissues
• Lipid Triad in diabetics: elevated TAGs, low HDL, small dense LDLs