biochemi Vol 6

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Questions and Answers

What is the primary consequence of the genetic mutation in Liddle syndrome?

Increased secretion of urea

Excessive reabsorption of Na+ and water (correct)

Increased production of aldosterone

Decreased ENaC activity

Which mechanism allows the removal of ENaC from the cell membrane in normal conditions?

Phosphorylation of ENaC

Ubiquitination by Nedd4-2 (correct)

Enzymatic digestion by proteolytic enzymes

Hydrolysis of sodium

How does increased ENaC activity contribute to hypertension?

By reducing blood plasma volume

By increasing sodium and water retention (correct)

By enhancing renal filtration rate

By elevating urea concentration in blood

What is the main role of alanine (Ala) in the glucose-alanine cycle?

Transport of nitrogen from muscle to liver Signup and view all the answers

What happens to excess NH4+ produced in tissues?

It combines with Glu to form Gln Signup and view all the answers

Which of the following correctly describes the process of transamination?

Transfer of an amino group to form an α-keto acid Signup and view all the answers

During the hepatic processing of alanine, what does it convert back into after donating its amino group?

Pyruvate Signup and view all the answers

What is the end product of oxidative deamination of glutamate in the liver?

α-Ketoglutarate Signup and view all the answers

What initiates the formation of Gln from Glu and NH4+?

Phosphorylation of Glu Signup and view all the answers

Which of the following statements about the epithelial sodium channel (ENaC) is NOT true?

ENaC activity is unaffected by hormonally regulated pathways. Signup and view all the answers

What role does PLP (Pyridoxal phosphate) play in the transamination process?

It serves as a cofactor for Aminotransferase. Signup and view all the answers

Which of the following statements accurately describes the Ping-pong reaction mechanism?

First product is released before the second substrate is attached. Signup and view all the answers

Which intermediate is formed during the transamination process involving PLP?

Pyridoxamine phosphate Signup and view all the answers

What happens to the concentration of GOT and GPT in the bloodstream when liver cells are damaged?

They increase, indicating liver damage. Signup and view all the answers

What does the reaction mechanism involving PLP utilize to facilitate the transfer of amino groups?

Covalent bonding through Schiff base formation. Signup and view all the answers

Why is the Schiff base formed during transamination considered unstable?

It can disassociate and reform with other substrates. Signup and view all the answers

What key function does PLP perform aside from its role in transamination?

It facilitates the synthesis of neurotransmitters. Signup and view all the answers

Which of the following accurately describes the general process of transdeamination?

Combines transamination and oxidative deamination. Signup and view all the answers

Which compound is primarily formed from the reaction of α-Ketoglutarate and Pyridoxamine phosphate?

L-Glutamate Signup and view all the answers

What implication does the function of ALT (GPT) in the body serve?

It reflects liver inflammation. Signup and view all the answers

What hormone is secreted by enteroendocrine cells in the duodenum in response to fatty acids and amino acids?

Cholecystokinin (CCK) Signup and view all the answers

What is the function of enteropeptidase in relation to trypsinogen?

It activates trypsinogen to trypsin. Signup and view all the answers

Which proteolytic enzyme cuts at the C-terminus of a polypeptide?

Carboxypeptidase Signup and view all the answers

What is the primary mechanism for intracellular protein degradation?

Proteasomal degradation Signup and view all the answers

Which enzyme is responsible for transferring the ubiquitin to the target protein?

E3 Signup and view all the answers

What does the presence of ubiquitin on a protein signify?

The protein is marked for degradation. Signup and view all the answers

What is the function of the ubiquitin-activating enzyme (E1)?

It activates ubiquitin. Signup and view all the answers

Which part of the ubiquitination process is primarily responsible for identifying the target protein?

E3 Signup and view all the answers

During the ubiquitin-proteasome pathway, what happens to the ubiquitin tag once the target protein is degraded?

It is reused. Signup and view all the answers

Which of the following accurately describes the role of aminoproteinases?

They cleave only at the N-terminus. Signup and view all the answers

What is the role of N-Acetylglutamate in the urea cycle?

It activates Carbamoyl phosphate synthetase I. Signup and view all the answers

Which genetic disorder is associated with a deficiency of Ornithine transcarbamoylase?

Hyperammonemia type 2 Signup and view all the answers

What causes the accumulation of N-Acetylglutamate in Hyperammonemia/NAGS deficiency?

Insufficient synthesis of N-Acetylglutamate. Signup and view all the answers

Which condition is characterized by the inability to convert Argininosuccinate to Arginine?

Argininosuccinicaciduria Signup and view all the answers

What is the consequence of Ornithine permease dysfunction in HHH syndrome?

Increased production of NH4+. Signup and view all the answers

Which medication utilizes Glycine to assist in the excretion of NH4+?

Benzoate Signup and view all the answers

What is the primary consequence of Arginase deficiency?

Increased levels of Ornithine. Signup and view all the answers

What compound is formed when Carbamoyl phosphate reacts with Ornithine?

Citrulline Signup and view all the answers

How does Phenylbutyrate help in NH4+ excretion?

It modifies Gln for urinary excretion. Signup and view all the answers

What condition is associated with an elevated Km value for Argininosuccinate synthetase?

Citrullinemia Signup and view all the answers

What role does leghemoglobin play in the relationship between legumes and nitrogen-fixing bacteria?

It absorbs oxygen to create a low-oxygen environment. Signup and view all the answers

Which statement best describes positive nitrogen balance?

It happens when nitrogen intake exceeds nitrogen output. Signup and view all the answers

What is a common symptom of Kwashiorkor disorder?

Fluid retention in soft tissues. Signup and view all the answers

Which enzyme complex is primarily involved in the nitrogen fixation process?

Nitrogenase complex. Signup and view all the answers

Which amino acids must be obtained through diet as they cannot be synthesized by the body?

Lysine and Tryptophan. Signup and view all the answers

What typically causes a negative nitrogen balance?

Excessive exercise without protein intake. Signup and view all the answers

Which factor is NOT relevant to the protein turnover rate in adults?

Diets low in carbohydrates. Signup and view all the answers

What happens to interstitial fluid in the human body?

It is constantly exchanged with blood vessels. Signup and view all the answers

In the nitrogen fixation process, what are pyruvate converted into?

Acetyl-CoA. Signup and view all the answers

What is a potential consequence of a lack of essential amino acids?

Inhibition of protein synthesis. Signup and view all the answers

Study Notes

Introduction

Legumes and nitrogen-fixing bacteria symbiosis is mutually beneficial
Legumes provide Leghemoglobin, which absorbs oxygen to create a low oxygen environment that enables nitrogenase activity and a substantial amount of oxygen for bacterial aerobic respiration.
They also provide bacteria with inorganic salts and organic nutrients.

Nitrogen fixation

Nitrogenase complex uses 4 pyruvate molecules that are converted into 4 acetyl-CoA.
The 8 electrons produced are transferred to N2 via ferredoxin or flavodoxin, which are then transferred to dinitrogenase reductase and dinitrogenase.
Dinitrogenase and dinitrogenase reductase are oxygen-phobic enzymes.
Leghemoglobin is crucial for absorbing oxygen and creating a low oxygen environment.

Nitrogen Balance

Normal adults maintain dynamic nitrogen balance by ingesting and excreting the same amount of nitrogen.
Positive nitrogen balance exists when intake exceeds excretion.
- Examples include growth phases in children, pregnancy, and recovery from illness.
Negative nitrogen balance exists when intake is less than excretion.
- Examples include surgery, cancer, Kwashiorkor disorder (high carbohydrate, low protein diet), and Marasmus disorder (lack of calories and specific amino acids).

Non-Essential and Essential Amino Acids

Non-essential amino acids can be synthesized within the body by enzymes.
Essential amino acids cannot be synthesized internally and rely on food intake for acquisition.
Conditionally essential amino acids can potentially be synthesized by the body, but the process is complex and may not be sufficient for rapid growth.
- Examples include Arg, Cys, Gln, Tyr, Gly, Pro, Ser, and Ornithine.
Lack of essential amino acids causes imbalances, and the deficient amino acid becomes the limiting factor for protein synthesis.

Reasons for Adult Protein intake

Supplementing depleted cells and enzymes such as digestive enzymes, gastrointestinal cells, skin cells, red blood cells, and antibodies.
Maintaining a rapid protein turnover rate, with approximately 500 grams generated and only 100 grams consumed daily, meaning 80% is synthesized from degraded proteins.

Kwashiorkor disorder

Occurs in children after weaning who consume high levels of carbohydrates but lack sufficient protein.
Symptoms include fatigue, restlessness, easy fatigue, and developmental delays, muscle loss, edemas, abdominal distension, and weakened immunity.
Common in children from countries experiencing droughts and famines.
Edema results from insufficient protein intake, leading to inadequate albumin production for blood osmotic pressure, leading to fluid retention in tissues.

Hydrostatic pressure and oncotic pressure

Fluid balance between blood vessels and tissues

Interstitial fluid needs constant renewal.
Blood vessels transport the fluid to organs like the lungs and kidneys for exchange, requiring a proper molecular and water balance to maintain internal homeostasis.
Water movement is controlled by the pressure difference between the tissues and the blood vessels.

Intestines

When fatty acids and amino acids enter the duodenum, they stimulate intestinal cells to secrete cholecystokinin (CCK), a peptide hormone that promotes digestive enzyme secretion from the pancreas and bile release from the gallbladder.
The pancreas releases trypsinogen, chymotrypsinogen, procarboxypeptidase A, and procarboxypeptidase B.
Trypsinogen is activated by enteropeptidase into trypsin, which activates chymotrypsinogen and procarboxypeptidase.
Proteases that cleave peptides include carboxypeptidase (cleaves C-terminus), aminopeptidase (cleaves N-terminus), and enteropeptidase (cleaves peptide chain middle).
Digested and absorbed amino acids are transported to the liver via the hepatic portal vein.

Intracellular protein degradation

Intracellular protein degradation primarily occurs through proteasomes and lysosomes, typically involving ubiquitination as a hydrolytic tag.
Ubiquitination steps:
- Activation: E1 ubiquitin activating enzyme receives and activates ubiquitin.
- Conjugation: E1 transfers the activated ubiquitin to ubiquitin conjugating enzyme E2.
- Recognition and ligation: E3 ubiquitin ligase transfers the E2-ubiquitin complex to the target protein, specifically recognizing the substrate and binding to E2 to transfer ubiquitin to the protein.
- Analogy: ubiquitin is like a tape (label attached to the target protein), E1 is like a tape dispenser (initial binding point of the tape), E2 is like holding the tape (transfer to the hand), and E3 is like a decision-making mind (identifies and sticks it).
E3 is the main regulator controlling the ubiquitination method and location.

Liddle syndrome

An autosomal dominant genetic disease caused by mutations in the epithelial sodium channel (ENaC) that prevent proper ubiquitination-mediated degradation.
This results in excess ENaC, leading to increased Na+ and water influx, causing hypertension.
60% of the human body is water: 40% intracellular, 20% extracellular (16% interstitial fluid, 4% blood), used for transporting nutrients and waste.
The kidneys are responsible for eliminating excess water. If there are issues with excretion, such as excessive NaCl absorption, it leads to water retention and hypertension.
The epithelial sodium channel (ENaC) is responsible for Na+ reabsorption in the distal convoluted tubule.
By regulating the amount of ENaC, it controls water entry and exit. Excessive ENaC leads to excess Na+ reabsorption, Cl- enters to maintain electrical neutrality, and consequently, water enters the blood, causing hypertension.
Nedd4 provides ubiquitin tags and removes ENaC. Normal ENaC interacts with Nedd4-2 in two ways: when ENaC mutates, rendering it un-taggable by ubiquitin from Nedd4-2, both mechanisms fail.
- Monoubiquitination (one ubiquitin attached to different amino acids): targets the protein to the lysosome for degradation.
- Polyubiquitination (multiple ubiquitins attached to the same amino acid): targets the protein to the proteasome for degradation.

Glucose-alanine cycle

The glucose-alanine cycle transports NH4+ from skeletal muscle to the liver.
1. Skeletal muscle:
- Proteins are degraded into amino acids.
- Transamination 1: the amino group is transferred to alpha-ketoglutarate, forming glutamate.
- Transamination 2: Glutamate transfers the amino group to pyruvate, forming alanine.
- Alanine carries the amino group to the liver via the bloodstream.
1. Liver:
- Alanine transfers its amino group to alpha-ketoglutarate, transforming itself back to pyruvate (can undergo gluconeogenesis).
- Glutamate is deaminated (oxidative deamination) back to alpha-ketoglutarate, continuing to receive other amino groups.
- The removed amino group is eventually metabolized into urea.
1. The mechanism of transferring the amino group of amino acids in skeletal muscle to alanine (transamination):
- Amino acids react with alpha-ketoglutarate to generate an alpha-keto acid and L-glutamate. This reaction is catalyzed by aminotransferase, with PLP acting as a coenzyme.
- L-glutamate reacts with pyruvate in skeletal muscle to form alpha-ketoglutarate and alanine, catalyzed by alanine aminotransferase.
1. Functions of alanine:
- Transport pyruvate (a high-energy, three-carbon compound) to the liver as a new energy source.
- Transfer nitrogenous waste from muscle to the liver for urea formation and excretion.

Glutamine as a carrier for NH4+ in blood

In many tissues, NH4+ produced from amino acid or nucleotide breakdown is toxic, thus combining with glutamate to form glutamine, which is then transported by blood to the liver.
Steps:
- 1. Phosphorylation of glutamate: Glutamate is phosphorylated by glutamine synthetase, consuming 1 ATP to form gamma-glutamyl phosphate.
- 1. Glutamine formation: gamma-glutamyl phosphate reacts with NH4+ in tissues, catalyzed by glutamine synthetase, releasing the phosphate group.
- 1. Glutamine is transported to the liver by blood.
The liver is responsible for metabolizing nitrogenous waste, and the burden of muscle metabolic products is transferred to the liver. Hence the saying, "If the liver is not good, the body is not good!"

From amino acids to urea: three key steps

Transamination

All amino acids undergo transamination to form L-glutamate for amino group metabolism.
Amino acids and alpha-ketoglutarate react with aminotransferase and PLP to produce alpha-keto acid and L-glutamate.
PLP (pyridoxal phosphate, the active form of vitamin B6) is a cofactor for all aminotransferases as its C=O group readily attaches to amino groups to form an imine. It serves as a covalent-bonded carrier for alpha-amino groups.
PLP binds to the Lys residue of aminotransferase via a Schiff base linkage (containing an imine), which is unstable and easily breaks in alkaline environments, forming a new Schiff base linkage with other substances.

PLP as coenzyme reaction mechanism

PLP binds to alpha-amino groups of amino acids, forming new Schiff bases, which are unstable and easily undergo a reverse reaction to form new Schiff bases with similar molecules, utilizing this property for transamination.
The aldehyde group on PLP acts like a baseball glove that can catch an amino group (ball) and then pass it on.
The amino acid residue on transaminase acts as a base pulling off the hydrogen from carbon, and then attaching a hydrogen in another position. This shifts the position of C=N.
- Before the C=N shift, the imine is formed as a ketone (ketone and imine are considered synonymous) by PLP acting as a ketone and the amino acid acting as the amino group donor.
- After the C=N shift, the imine is formed as an amine by PLP acting as the amino group donor and the amino acid acting as the ketone. This imine is then hydrolyzed, successfully transferring the amino group to PLP.
Through a series of electron transfers, amino acids form alpha-keto acids that leave, and PLP becomes pyridoxamine phosphate.
Alpha-ketoglutarate reacts with pyridoxamine phosphate, accepting the alpha-amino group and creating glutamate. This is the reverse reaction of the previous steps, but instead of the original product, alpha-keto acid, a different alpha-keto acid (alpha-ketoglutarate) reacts, reversing the steps once.

Ping-pong reaction

The enzyme retains a portion of the first substrate through covalent bonds and releases the first product before accepting the second substrate. This can be shown as:
- E + S1 ↔ ES1 ↔ E’P1 ↔ P1 + E’
- E’ + S2 ↔ E’S2 → E + P2
PLP reaction mechanism can be considered a Ping-pong reaction:
- PLP reacts with amino acids to produce alpha-keto acids and pyridoxamine phosphate.
- The alpha-keto acid leaves.
- Alpha-ketoglutarate is added and reacts with pyridoxamine phosphate to produce L-glutamate.
Given that most of these steps are reversible, the overall reaction can be considered as proceeding forward, releasing the first product, accepting the second substrate, and then proceeding backward.

GOT, GPT as Indicators of Liver damage

GOT and GPT are enzymes in the liver. When liver cells are damaged or injured, the concentration of GOT and GPT in the blood increases, serving as an indicator of liver damage.

Other PLP functions

PLP, besides its role in transamination, has two other notable functions: Arginase and the biosynthesis of porphyrin precursors.

Short-term regulation

When consuming a high-protein diet, amino acid levels in the body increase. Glutamine, one of the amino acids, reacts with acetyl-CoA in the presence of N-acetylglutamate synthase (NAGS) to form N-acetylglutamate.
This substance activates carbamoyl phosphate synthetase I (the first enzyme in the urea cycle), promoting urea synthesis.

Genetic diseases associated with the urea cycle

Examining the flow chart of the cycle allows for understanding the disease process and connection between the diagram and disease names.

Hyperammonemia type 1

Hyper- means super/over, and -emia means associated with blood.
Carbamoyl phosphate synthetase I is deficient or mutated, losing function.
The affected reaction is NH4+ + HCO3- + 2 ATP → Carbamoyl phosphate + 2 ADP + Pi.

Hyperammonemia type 2

An X-linked recessive genetic disease involving Ornithine transcarbamoylase.
The affected reaction is: Carbamoyl phosphate + ornithine → Citrulline.

Hyperammonemia/NAGS deficiency

Deficiency in N-acetylglutamate synthase leads to insufficient production of N-acetylglutamate, preventing the activation of carbamoyl phosphate synthetase I (the first enzyme in the urea cycle).

HHH (Hyperornithinemia, Hyperammonemia, Homocitrullinuria) syndrome

Ornithine permease transports synthesized ornithine from cytosol to mitochondria.
Dysfunctional ornithine permease causes ornithine accumulation (hyperornithinemia), which also leads to NH4+ accumulation (hyperammonemia).
-uria means urine content.
In this scenario, there's no ornithine in the mitochondria to react with carbamoyl phosphate, so carbamoyl phosphate reacts with lysine, forming homocitrulline.
Normally, urine does not contain this compound, so the presence of excess homocitrulline in urine (homocitrullinuria) indicates potential health issues..

Citrullinemia

Deficiency in argininosuccinate synthetase.
Patients have levels of argininosuccinate synthetase too low to detect in cells, resulting in increased Km value compared to normal individuals (25 times higher), meaning argininosuccinate formation rate is much lower at the same citrulline concentration.

Argininosuccinicaciduria

Deficiency in argininosuccinase, leading to argininosuccinate accumulation.
The affected reaction is: Argininosuccinate → Arginine + Fumarate.

Hyperargininemia

A body autosomal recessive genetic disease resulting from low levels of arginase, causing elevated intracellular Arg levels.
The affected reaction is Arg hydrolysis producing urea and ornithine.

Treatment for genetic diseases associated with the urea cycle

Overview

When enzymes involved in the urea cycle become dysfunctional, NH4+ becomes difficult to eliminate.
Common treatments use benzoate and phenylbutyrate as medications to modify Gly and Gln, forming hippurate or phenylacetylglutamine, which can be excreted in urine, providing an alternative pathway for NH4+ elimination.

Mechanism of action

Gly and Gln are coupled to the drugs with the help of CoA-SH.
The two modified amino acids are not reabsorbed by renal tubules and are excreted in urine.
Reabsorption of amino acids typically requires carrier proteins, which can't recognize the modified amino acids.
Normally, the kidneys excrete waste products first, and then reabsorb essential substances like amino acids to prevent potential loss. The drugs work by modifying amino acids, making them unrecognizable to carrier proteins, resulting in excretion.

Medications

Benzoate:
- It forms benzoyl-CoA with CoA-SH, then reacts with Gly to form hippurate, which is excreted in urine.
Phenylbutyrate:
- It forms phenylacetyl-CoA with CoA-SH, then reacts with Gln to form phenylacetylglutamine, which is excreted in urine.

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