Biochem - Proteins, AA Catabolism vs Urea Cycle

Question 1

Conventionally a peptide chain is written in what direction?

Answer 1

The opposite as it is synthesized, starting with the N terminus

Synthesis starts with the C-terminus

Question 2

Cathode and Anode in electrophoresis

Answer 2

Chathode (-)

Anode (+)

Question 3

Alpha Helix

Answer 3

-NH H-bond every four AA’s along the chain

Question 4

Beta Sheet

Answer 4

H-bond between C=O and -NH

Question 5

Secondary Stx Motifs

Answer 5

Helix-turn-helix

Leucine zipper

Zinc finger

Found within transcription factors

Question 6

Structural subunit of F-actin

Answer 6

Oligomer

Question 7

Enthalpy of protein folding

Answer 7

Essentially isothermic (change in enthalpy is very small)

Although many H-bonds are broken during protein formation, the same number are broken from water

Question 8

What energetically drives protein folding?

Answer 8

Entropy - resulting from increased disorder of water molecules as hydrophobic regions associate inside the protein

Question 9

Molten globules

Answer 9

Intermediate between secondary structures and the folded protein

Question 10

Heat shock proteins

Answer 10

Chaperones - prevent the aggregation of newly synthesized proteins by binding to their hydrophobic regions

I.e. HSP10, HSP 60, and HSP70

Question 11

Myoglobin vs. Hemoglobin

Answer 11

Myoglobin - single peptide with one O2 binding site

HbA - tetrameric with two alpha and two beta polypeptide chains (four binding sites) each conjugated with a heme group.

Question 12

R state

Hemoglobin

Answer 12

Relaxed state

When one O2 molecule binds to a heme of hemoglobin

Increases affinity of O2 binding

Positive cooperativity

Question 13

What reduces the O2 affinity of hemoglobin

Answer 13

Acidity (increased conc. of CO₂)

This is called the Bohr effect

Question 14

Binding of 2,3-bisphosphoglycerate (BPG)

Answer 14

Decreases the affinity of HbA for O2

BPG is by-product of glycolysis, this ensures efficient O2 offloading in tissues that are metabolically active, especially in high altitude or anemia, where intracellular BPG levels might be elevated

Question 15

Sickle-cell anemia

Answer 15

The beta chain of hemoglobin undergoes a point mutation that causes a missense (E6V) mutation in the reading frame

The introduction of hydrophobic valine promotes polymerization of deoxygenated mutant hemoglobin

HbS polymers adopt the classic “sickle” shape - prone to hemolysis leading to anemia

Question 16

Peptide configuration of collagen

Answer 16

Gly - X - Y

Gly - every third amino acid

X - typically proline

Y - typically hydroxyproline or hydroxy lysine

Collagen is a triple helical structure, where three polypeptide chains intertwine around each other

Question 17

Synthesis of collagen

Answer 17

pro-collagen syn. in rER, where the signal sequence (pre) is removed)

In the rER lumen, pro and lys residues undergo hydroxylation (depends on presence of oxygen and Vit C)

Hydroxylysine residues are further glycoslated with galactose and glucose

Modified peptide chains are assembled into the triple helix, forming procollagen, which is secreted outside the cell and processed to form mature collagen

Collagen fibers undergo cross-linking by oxidation of amino acid side chains

Question 18

EDS

Answer 18

Ehlers-Danlos Syndrome

Mutations in collagen synthesis and processing - fragile skin and hyper-elastic

Joint hypermobility

Question 19

Osteogenesis imperfect

Answer 19

Mutations in collagen I genes

Brittle bones which are prone to fracture

Question 20

Scurvy

Answer 20

Impairs the hydroxylation of proline and lysine residues

Results in unstable collagen (mp from 42 C to 24 C) as the result of the loss of interstrand H-bond fomration from lack of pro-OH

sxs - bleeding gums and poor wound healing

Question 21

Alport syndrome

Answer 21

Mutations in collagen IV disrupts the collagen network supporting kidney glomerular cells

Leads to compromised kidneys and failure

Question 22

Proteopathies

Answer 22

Diseases classified by misfolded proteins

Question 23

CJD

Answer 23

Creutzfeldt-Jakob Disease - average onset at 68 y/o

Progresses rapidly - death within year

Rare

Question 24

vCJD

Answer 24

Variant Creutzfeldt-Jakob Disease - younger-onset form, avg. death age of 28 y/o

Linked to consumption of beef from cows affected by bovine spongiform encephalopathy

Question 25

Kuru

Answer 25

Prion disease ID'd among the Fore tribe in Papua New Guinea ## Footnote Believed to have spread through cannabilistic practices

Question 26

PrP^c

Answer 26

Normal prion proteins - are primarily alpha-helical structures in the brain

Question 27

PrP^SC

Answer 27

Misfolded, disease-causing conformation of the prion, pirmarily beta-sheet conformation

Question 28

Propagation of prion disease

Answer 28

The misfolded PrP^SC acts as a template, causing endogenous PrP^C to also misfold

Question 29

Prion disease transmission

Answer 29

After consumption, PrP^SC crosses the intestinal lining, gets taken up by immune cells, and is transported to the brain

Question 30

Affect of prions disease in the brain

Answer 30

Accumulation of beta-sheet rich prions leads to the formation of protein plaques in the brain ## Footnote Hallmark of prion disease

Question 31

Alzheimer's

Answer 31

Buildup of misfoled Aβ42 protein in the brain APP undergoes enzymatic cleavages - resulting in Aβ42 fragments that can aggregate and form amyloid or senile plaques outside neurons Much like prions, Aβ42 plaques adopt a beta-sheet conformation, leading to the formation of insoluble aggregates (resists degradation through normal cellular processes) Intracellular tangles of tau (Aβ42 causes intracellular phosphorylation of tau protein) forms neurofibrillary tangles - believed to damage neurons from within ## Footnote Aβ42 originates from amyloid precursor protein (APP) - transmembrane protein believed to play role in neuron growth, survival, and repair post-injury

Question 32

Genetic factors of Alzheimer's disease

Answer 32

Mutations in the APP gene and componenets of the gamma-secretase enzyme (involved in APP cleavage) have been linked to increased Aβ42 production

Question 33

A patient with a history of recurrent vaso-occlusive crises is found to have a mutation in the β-chain of hemoglobin. Which of the following amino acid substitutions is most likely responsible for this patient's condition?

Answer 33

Glutamate to valine (E6V)

Question 34

What sequence of AA residues is most commonly found in collagen?

Answer 34

Gly-X-Y X - typically proline Y - either pro-OH or Lys-OH

Question 35

Difference between amyloid plaques and neurofirillary tangles?

Answer 35

Aβ42 plaques - outside of neurons Neurofibrillary tangles - inside of neurons (Aβ42 phosphorylates tau causing it to tangle)

Question 36

A deficiency in what vitamin impairs the hydroxylation of proline and lysine residues in collagen synthesis?

Answer 36

Vitamn C (Scurvy)

Question 37

Active site

Answer 37

Question 38

Michaelis-Mentin Eq

Answer 38

Question 38

K_m

Answer 38

Affinity of enzyme to substrate (lower K_m = higher affinity) Mathematically derived as the substrate concentration at which v = 1/2(v_max)

Question 39

When [S] <<< K_m ## Footnote How is v affected?

Answer 39

The rate is directly proportional to [S]

Question 40

When [S] >>> K_m

Answer 40

The rate approaches V_max and becomes independent of [S]

Question 41

Cofactors

Answer 41

Inorganic ions or organic molecules that bind to enzymes and are essential for activity Types: Posthetic groups and loosely bound cofactors Fx - assist substrate binding, stabilize enzyme stx, or participate directly in reaction by facilitating transfer of chemical groups or electrons

Question 42

Prosthetic Groups

Answer 42

Tightly and permanently bound to the enzyme (i.e. heme is a prosthetic group of hemoglobin) ## Footnote Type of cofactor

Question 43

Loosely Bound Cofactor

Answer 43

Can associate and dissociate from the enzyme (when bound, activate the enzyme)

Question 44

Coenzymes

Answer 44

Subset of cofactors that are organic molecules - often derived from vitamins Unlike cofactors, coenzymes are **not permanently bound** to the enzyme Fx - carriers, transporting specific groups or electrons from one reaction to another (many redox rx'ns) ## Footnote i.e. NAD+ and NADP+ (derived from niacin) CoA (derived from panthenic acid) FAD (derived from riboflavin)

Question 45

Affect of cofactors/coenzymes on enzyme

Answer 45

Catalytic activity Specificity Regulation (allosteric) Diversity of rx'ns

Question 46

Cooperativity

Answer 46

The binding of a substrate to one active site can affect the binding of substrates to other active sites

Question 47

Allosteric enzymes often display _____ kinetics when rx'n velocity is plotted against [S]

Answer 47

Sigmoidal

Question 48

Affects of allosteric regulation

Answer 48

1. Fine-tuning of metabolic pathways 2. Feedback inhibition 3. Rapid response 4. Integration of mlutiple signals

Question 49

Competitive inhibitors

Answer 49

Can be overcome by adding more substrate (increasing [S]) Ex: Statins competitively inhibit HMG-CoA reductase ## Footnote V_max remains unchanged K_m increases (a higher [S] is need to achieve half of V_max)

Question 50

Noncompetitive inhibitors

Answer 50

Bind to allosteric site, causing conformational change Increasing [S] does not overcome inhibition Ex: heavy metals like lead or mercury ## Footnote V_max decreases K_m remains unchanged (affinity is not affected)

Question 51

Lineweaver-Burke

Answer 51

Question 52

Enzyme inhibition graphs

Answer 52

Question 53

Graphs of V vs [S] for an allosteric enzyme

Answer 53

Does not follow MM kinetics, but is hyperbolic and the v vs. [S] is sigmoidal

Question 54

Inborn errors of metabolism

Answer 54

PKU, galactosemia, and Tay-Sachs disease

Question 55

Accumulation of Phe

Answer 55

PKU - deficiency of phenylalanine hydroxylas leads to accumulation of phenylalanine which causes intellectual disability if not managed

Question 56

G6PD deficiency

Answer 56

glucose-6-phosphate dehydrogenase deficiency can lead to hemolytic anemia (especially after exposure to certain drugs or infections)

Question 57

Disorder of purine and pyrimidine metabolism

Answer 57

Can lead to conditions like Lesch-Hyhan syndrome or orotic aciduria

Question 58

Congenital adrenal hyperplasia

Answer 58

An **endocrine disorder** that can result from deficiencies in enzymes involved in cortisol synthesis

Question 59

Where are AA's abosrbed

Answer 59

Intestinal epithelial cells and taken up into cellular AA pools

Question 60

Essential AA's

Answer 60

Cannot be made and must be obtained through diet PVT TIM HaLL (Phe, Val, Trp, Thr, Ile, Met, His, Lys, Leu) ## Footnote a = arginine (conditionally essential during periods of rapid growth)

Question 61

Primary reserve of AA's

Answer 61

Muscle (this includes reserve for essential AA's) Diets deficient in essential AA's can cause muscle wasting (when diet is deficient in essential AA's, it can take up to six weeks for sxs to manifest due to muscle reserves)

Question 62

Kwashiorkor malnutrition

Answer 62

Protein deficiency with adequate calories Children develop swollen belly (ascitic belly) becuase albumin is 97% of serum protein and is responsible for oncotic pressure ## Footnote Fluid collects in abdomen due to gravity

Question 63

Marasmus malnutrition

Answer 63

Inadequate energy intake in all forms, including protein

Question 64

Cystinuria

Answer 64

**Autosomal recessive** Impairs the reabsorption of dibasic AA (disulfide linked Cys-Cys, ornithine, arginine, and lysine) Hexagonal crystals of cystine in urinalyses are diagnostic of cystinuria Recurrent nephrolithiasis (kidney stones) ## Footnote Management: Low cystine diet and increased fluid intake Potassium citrate and sodium bicarb (make urine less acidic and increases solubility of cystine) Pencillamine and topronin are cystine-binding thiol drugs that bind and form a water-soluble complex

Question 65

Hartnup Disease

Answer 65

**Autosomal recessive** Affects renal tubule reabsorption of **neutral** AA's, importantly tryptophan Trp is used to synthesize B3 (niacin) so sxs resemble deficiency of niacin (pellagra) ## Footnote Management: Dietary intake of trp-rich foods, trp and niacin supplements

Question 66

3 D's of Pellagra

Answer 66

Dermatitis Diarrhea Dementia (Death) Other malabsorption disorders and chronic alcoholism can present with pellagra like sxs ## Footnote Corn-based diets (low in trp and niacin) increase risk of pellagra

Question 67

Hartnup vs. Cystinuria

Answer 67

Question 68

Ammonia toxicity

Answer 68

Toxic, especially to the CNS (causing encephalopathy) ## Footnote When ammonia levels are high, astroctyes convert excess into glutamine, which is osmotically active. This draws water into astrocytes leading to cellular swelling and cerebral edema

Question 69

Disposal of ammonia

Answer 69

80-90% of ammonia is disposed via **formation of urea in the liver** and excreted by the kidneys ## Footnote Carried in blood by alanine (muscle) and glutamine (most tissues)

Question 70

What are the most abundant AA's in the blood?

Answer 70

Alanine (muscle) and glutamine (most tissues) Transport ammonia to the liver

Question 71

Enzymes of AA catabolism

Answer 71

Amino transferases (aka transaminases) - transfer NH₂ from an AA to another alpha-keto acid, which is converted to its cognate AA Requires pyridoxal phosphate (PLP) a B6 derivative, as a cofactor Reversible to allow interconversion of AA and ketoacids as needed for metabolic need Specific aminotransferases for each AA (except thr. and lys) -Most use alpha-ketoglutarate and glutamate ## Footnote Thus amino groups are funneled i nto glutamate by transamination reactions

Question 72

ALT

Answer 72

Crucial in the glucose-alanine cycle (allows the body to transport nitrogen and carbon between muscles and the liver to **maintain blood glucose levels during fasting or exercise, providing a way to remove excess nitrogen from muscle tissue.** In muscle, when AA are broken down their amino groups are transferred by aminotransferases to alpha-KG to generate Glu. ALT then transfers the amino from Glu to pyruvate producing Ala. Ala is released into the blood and travels to the livers The liver converts Ala back to pyruvate and Glu The pyruvate can be used to produce glucose The amino group from glutamate is used to form urea

Question 73

AST

Answer 73

Aspartate transaminase

Question 74

Where are ALT and AST highly expressed?

Answer 74

Liver ## Footnote Also expressed in heart, muscle, and kidneys

Question 75

Glutamine Synthetase

Answer 75

Glu + NH₃ +ATP -> Gln + ADP + P_i GS fixes free NH₃ to glutamate (produced by transaminases) to make glutamine which is released into the bloodstream for transport to liver ## Footnote Kidneys and brain also take up Gln for metabolic use

Question 76

Glutaminase

Answer 76

Releases NH₃ from glutamine to regenerate glutamate In the liver, the released NH₃ enters the urea cycle Glutamate is deaminated again by glutamate DH which regenerates alpha-ketoglutarate

Question 77

Glutaminase in kidney

Answer 77

Releases NH₃ from glutamine to help regulate urine pH

Question 78

Glutaminase function in the brain

Answer 78

The formation of glutamate from glutamine via glutaminase is a pathway used to form the excitatory neurotransmitter (glutamate)

Question 79

Glutamate DH

Answer 79

Deaminates glutamate to form alpha-ketoglutarate/NAD(P)H/NH₃ ## Footnote Reversible rx'n that is used to fix free ammonia to form glutamate

Question 80

Why is glutamate unique?

Answer 80

Only AA that undergoes rapid oxidative deamination

Question 81

In the liver, what enzymes release ammonia to enter the urea cycle?

Answer 81

Glutaminase and glutamate DH

Question 82

Carbomyl Phosphate Synthetase I

Answer 82

Step 1 of urea cycle Rate-limiting step N-acetyl glutamate is essential allosteric activator of CPS-I (formed by NAG synthetase which is activated by R) ## Footnote Mitochondrial enzyme

Question 83

Why does adding arginine dietary intake help Tx Pt's with some urea cycle disorders?

Answer 83

Arginine activates N-acetyl glutamate synthetase, which allosterically activates CPS-I

Question 84

Ornithine Transcarbamoylase (OTC)

Answer 84

Step two of urea cycle carbamoyl phosphate + ornithine -> citrulline ## Footnote Mitochondrial enzyme

Question 85

Argininosuccinate Synthetase (AS)

Answer 85

Step three or urea cycle citrulline + aspartate -> argininosuccinate ## Footnote AST supplies aspartate for this step

Question 86

Argininosuccinate lyase

Answer 86

Step four of urea cycle argininosuccinate -> arginine + fumarate

Question 87

Arginase

Answer 87

Step five of urea cycle arginine -> urea + ornithine

Question 88

Hyperammonemia

Answer 88

Causes: Liver dysfunction Urea cycle disorders (UCD) Sxs: asterixis (flapping tremor), vomiting, confusion, blurred vision, slurred speech, seizures

Question 89

Only effective means to rapidly reduce level of circulating ammonia

Answer 89

Hemodialysis

Question 90

Ammonia scavengers

Answer 90

Form water soluble products with gly or glu that are excreted by the kidneys, providing alternate pathway for removal of NH₃ **Sodium benzoate** - combines with gly in liver to form hippuric acid which is excreted **Sodium phenylbutyrate** - metabolized to phenylacetate which reacts with gln to form phenylacetylglutamine which is excreted

Question 91

Lactulose

Answer 91

Non-absorbable disaccharide used in context of liver diseases. Fermentation of lactulose by gut bacteria lowers colonic pH. Lower colonic pH promotes conversion of ammonia to ammonium, which do not easily diffuse across gut epithelium ## Footnote NH₃ is then able to be excreted in feces

Question 92

Rifaximin

Answer 92

Abx used to reduce the population of ammonia-producing bacteria in the gut

Question 93

Long-term management of hyperammonemia

Answer 93

Low-protein diet Ammonia scavenger meds Liver transplantation

Question 94

Urea cycle disorders

Answer 94

A complete lack of any enzyme of the urea cycle results in death shortly after birth Deficiencies present after first feeding (first protein rich meal) Deficiency of urea cycle enzyme: Increased serum levels of gln

Question 95

CPS-I deficieCPSD)

Answer 95

Aka hyperammonemia-I Most severe UCD

Question 96

A 30-year-old woman is concerned about her pregnancy after learning that her brother has a urea cycle disorder. She is currently 10 weeks pregnant and asymptomatic. Which of the following tests would be most appropriate to screen for the carrier status of ornithine transcarbamylase deficiency in this patient? 1. Serum ammonia level 2. Urinary orotic acid after protein load 3. Plasma citrulline level 4. Liver biopsy 5. Serum urea nitrogen level

Answer 96

**Urinary orotic acid after protein load.** Rationale: This test is most appropriate for screening for carrier status of ornithine transcarbamylase (OTC) deficiency. OTC deficiency is an X-linked disorder, and female carriers can be asymptomatic. The protein load test stimulates the urea cycle, and in OTC deficiency carriers, this leads to increased orotic acid excretion due to the accumulation of carbamoyl phosphate, which is then shunted into the pyrimidine synthesis pathway.

Question 97

A 28-year-old man presents to the clinic with a history of recurrent skin rashes, particularly on sun-exposed areas. He also complains of intermittent diarrhea and occasional confusion. Physical examination reveals a rough, scaly rash on face and hands. Which of the following amino acids is most likely affected in this condition? 1. Phenylalanine 2. Tryptophan 3. Leucine 4. Methionine 5. Arginine

Answer 97

**Tryptophan** Rationale: The clinical presentation of recurrent skin rashes on sun-exposed areas, intermittent diarrhea, and occasional confusion is consistent with Hartnup disease. Hartnup disease is a disorder of neutral amino acid transport, particularly affecting tryptophan absorption. Tryptophan is a precursor for niacin (vitamin B3), and its deficiency leads to pellagra-like symptoms, which include dermatitis, diarrhea, and dementia (the "3 D's" of pellagra).

Question 98

A newborn presents with lethargy, poor feeding, and respiratory distress within 48 hours of birth. Laboratory tests reveal elevated plasma ammonia and trace amounts of citrulline. Urinary orotic acid levels are normal. Which of the following is the most likely diagnosis? 1. Ornithine transcarbamylase deficiency 2. Argininosuccinate synthetase deficiency 3. Carbomoyl phosphate synthetase I deficiency 4. Argininosuccinate lyase deficiency 5. N-acetylglutamate synthase deficiency

Answer 98

**Carbamoyl phosphate synthetase I deficiency** Rationale: The presentation of hyperammonemia within 48 hours of birth, along with trace amounts of citrulline and normal urinary orotic acid levels, is consistent with carbamoyl phosphate synthetase I (CPS-I) deficiency. CPS-I catalyzes the first step of the urea cycle. Its deficiency leads to ammonia accumulation and very low or absent citrulline. Normal orotic acid levels distinguish it from ornithine transcarbamylase deficiency, which would show elevated orotic acid.

Question 99

A 45-year-old man with a history of chronic alcoholism presents with confusion, asterixis, and a "sweet" odor on his breath. His blood ammonia level is 180 μmol/L. Which of the following medications would be most appropriate for immediate management? 1. Sodium benzoate 2. Rifaximin 3. Lactulose 4. Carbaglue (carglumic acid) 5. Penicillamine

Answer 99

** Lactulose** Rationale: The patient presents with signs of hepatic encephalopathy secondary to chronic alcoholism. Lactulose is the first-line treatment for acute management of hepatic encephalopathy. It works by lowering colonic pH, which promotes the conversion of NH3 to NH4+ ions, reducing ammonia absorption from the gut. The "sweet" odor on the breath is likely fetor hepaticus, often associated with severe liver disease.

Question 100

A patient with recurrent kidney stones is diagnosed with cystinuria. Which of the following is the primary defect in this condition?

Answer 100

**Impaired reabsorption of dibasic amino acids in the kidneys** Rationale: Cystinuria is characterized by impaired reabsorption of dibasic amino acids (cystine, ornithine, arginine, and lysine) in the kidneys. This leads to excessive excretion of these amino acids, particularly cystine, which has low solubility and tends to form kidney stones. The primary defect is in the transport system for these amino acids in the renal tubules, not in their metabolism or production.

Question 101

Drug company develops a therapy to treat a cell membrane protein misfolding disorder. This mutation leads to abnormal protein folding and subsequent intracellular degradation of the protein before it can reach the cell membrane. With the new combination of drug therapy, the first drug corrects the processing and trafficking of the protein, enabling it to reach the cell surface membrane. Once the protein has reached the cell surface, its function is enhanced by the second drug. This therapy is most likely to be helpful in which of the following conditions?

Answer 101

Cystic fibrosis

Question 102

Atrophy of CNS is common in many pathologies. What type of CNS atrophy is unique to Creutzfeldt-Jakob disease?

Answer 102

Atrophy of the cerebellum

Question 103

Steps of the urea cycle

Answer 103

Remember enzymes by: COAAA

Question 104

Specific sxs of osteogenesis imperfecta

Answer 104

Transulcent skin showing visible underlying veins Frequent bone fractures | Col I deficiency

Question 105

Specfic sxs of scurvy

Answer 105

Bleeding from gums

Question 106

Activator of NAG

Answer 106

Arginine ## Footnote NAG is an obligate activator of CPS I

Question 107

OTC deficiency inheritance pattern

Answer 107

X-linked recessive ## Footnote Only x-linked disease in the urea cycle Female carriers report aversion to high-protein foods (due to lionization of x-chromosome)