Session 5: Protein Metabolism Flashcards
Where does stage 1 of protein catabolism take place?
Extracellular: GI tract
What enzymes in the GI tract hydrolyse proteins in stage 1 of protein catabolism?
Proteases including peptidases hydrolyse the peptide bonds to release free amino acids
How are most free amino acids taken up into enterocytes?
Sodium-dependent secondary active transport by solute carrier proteins (SLC)
Free amino acids are released from enterocytes into the portal system and travel in the blood to other tissues where they are used for…
1) Protein synthesis = around ~75% proteins used for this
2) Synthesis of N-containing compounds = (purines, creatine, haem)
3) Oxidation = to produce energy (ATP)
What two hormones stimulate the uptake of amino acids into tissues such as skeletal muscle, adipose tissue and liver?
Insulin and growth hormone (GH)
What are the two mechanisms of protein catabolism?
Transamination, deamination
How many different types of variable side chain can arise in amino acids?
20
What are the 9 essential amino acids?
Isoleucine, Lysine, Threonine, Histidine, Leucine, Methionine, Phenylalanine, Tryptophan, Valine
Creatinine is a useful clinical marker of…
Renal function - increased levels in blood indicate damage to nephrons of the kidney
Creatinine urine excretion over 24h is proportional to ___ mass (estimate of ___ mass)
Creatinine urine excretion over 24h is proportional to muscle mass (estimate of muscle mass)
What is creatinine?
Product of muscle breakdown of creatine and creatine phosphate, measurement of renal function
What is transamination?
The transfer of an amino group from one amino acid to a keto acid, producing a new nonessential amino acid and a new keto acid
When does mobilisation of protein reserves take place?
During extreme stress (starvation)
In what disease does excessive breakdown of protein occur?
Cushing’s syndrome (excess cortisol)
Striae of the skin due to excessive protein breakdown causing weaknesses in the skin
Insulin and growth hormone stimulates ___ synthesis
Insulin and growth hormone stimulates protein synthesis
___ (e.g., cortisol) decreases protein synthesis
Glucocorticoids (e.g., cortisol) decreases protein synthesis
What is the name of the enzyme involved in transamination?
Aminotransferases
Aminotransferase enzymes require the coenzyme pyridoxal phosphate - a derivative of vitamin ___
Aminotransferase enzymes require the coenzyme pyridoxal phosphate - a derivative of vitamin B6
What are two important diagnostic markers in the blood test to indicate liver function?
Plasma alanine aminotransferase (ALT) and plasma aspartate aminotransferase (AST)
Plasma ALT and AST at heightened levels in the blood in conditions that cause extensive cellular necrosis such as…
Viral hepatitis, autoimmune liver diseases, toxic injury (death cap mushroom)
Where does deamination mainly occur?
Liver and kidney
At what range does ammonia need to be kept in the blood to be considered safe?
25-40 umol/L
Where does the urea cycle take place?
Occurs in the liver
- A high protein diet activates enzymes
- A low protein diet inhibits enzymes (refeeding syndrome)
What is refeeding syndrome?
Can occur when nutritional support is suddenly given to those who are severely malnourished patients e.g., anorexic patients
What are the symptoms of ornithine transcarbamylase (OCT) deficiency?
Lethargy → Ammonia is neurotoxic, depressing brain function.
Confusion → Ammonia disrupts neurotransmitters and causes cerebral edema.
Vomiting → The brainstem’s vomiting center is triggered by toxins.
Poor feeding → The body naturally avoids protein to reduce ammonia production.
Rapid breathing (Respiratory alkalosis) → The body hyperventilates to remove excess carbon dioxide and balance blood pH.
Seizures and coma → Severe ammonia toxicity leads to brain swelling and dysfunction.
What are the risk factors for refeeding syndrome?
BMI <16
Unintentional weight loss >15% in 3-6 months
10 days or more with little/no nutritional intake
Management of ornithine transcarbamylase (OCT) deficiency?
Low protein diet, replace amino acids in the diet with keto-acids
What test is used for diagnosing a number of paediatric diseases?
Heel prick test
What is the most common defect/enzyme deficiency in the urea cycle?
Ornithine transcarbamylase deficiency (OTC deficiency)
How many inherited diseases of amino acid metabolism does the heel prick test screen for?
> 50 inherited diseases involving defects in amino acid metabolism
What is phenylketonuria (PKU)?
Most common inborn error of amino acid metabolism (~1 in 15,000 births)
Deficiency in phenylalanine hydroxylase (PAH) enzyme
What does phenylketonuria (PKU) cause?
Accumulation of phenylalanine in tissue, plasma and urine.
Phenylketones in urine
What causes Phenylketonuria (PKU)?
Autosomal recessive condition (chromosome 12) leading to deficiency of phenylalanine hydroxylase (PAH)
What is the treatment of phenylketonuria (PKU)?
Low-phenylalanine diet + tyrosine supplementation
Avoid high protein foods
What are the symptoms of phenylketonuria (PKU)?
Severe intellectual disability, developmental delay, microcephaly, seizures, hypopigmentation
What is the most common cause of homocystinurias?
Defect in cystathionine beta-synthase (CBS) is most common
Defect in methionine synthase is also possible
What is homocystinuria?
Autosomal recessive disorder leading to excess homocysteine (oxidised form of homocysteine) excreted in urine
What causes disease symptoms in homocystinuria?
Accumulation of homocysteine and methionine
What is treatment of homocystinuria?
Low-methionine diet
Avoiding milk, meat, fish, cheese, eggs
Cysteine, vitamin B6, Betaine, B12 and folate supplementation
What are the symptoms of Homocystinuria?
Marfan-like body stature (long limbs and fingers), dislocation of the lens, intellectual disability
Elevated plasma homocysteine shown to be associated with ___ disease
Elevated plasma homocysteine shown to be associated with cardiovascular disease
What are some key features of urea?
High nitrogen content, non-toxic, extremely water-soluble, most urea excreted in urine via kidneys
Where is urea synthesised by the urea cycle?
Liver
Which enzymes can deaminate amino acids?
- Amino acid oxidases
- Glutaminase
- Glutamate dehydrogenase
What is deamination?
Removal of an amino group - removes ammonia (NH3)
What are major nitrogen containing compounds in the body?
Proteins, amino acids, purines & pyrimidines (DNA/RNA)
What are minor nitrogen containing compounds in the body?
Porphyrins (haem), creatine, neurotransmitters (e.g., dopamine), some hormones (e.g., adrenaline)
What are the conditionally essential amino acids?
Cysteine, Arginine, Tyrosine (CAT)
What is the reference range of average excretion of creatine in urine per day for men and women?
Men = 14-26 mg/kg
Women = 11-20 mg/kg
All 20 amino acids are required for ___ synthesis
All 20 amino acids are required for protein synthesis
Phenylketonuria (PKU)
- Autosomal recessive
- Deficiency in phenylalanine hydroxylase (PAH) enzyme
- Accumulation of phenylalanine in tissue, plasma, urine
- Phenylketones in urine
- Phenylketones oxidised to phenylacetate = musty smell excreted in urine
Symptoms of phenylketonuria
Severe intellectual disability, developmental delay, microcephaly, seizures, hypopigmentation
Treatment of phenylketonuria
- Low phenylalanine diet
- Supplement with tyrosine
- Avoid artificial sweeteners containing phenylalanine
- Avoid high protein food (milk, meat, egg)
Diagnosis of phenylketonuria
- Detection of phenylketones in the urine
- Elevated blood serum phenylalanine concentration [normally less than 0.1mmol/L; in PKU can exceed >1.0mmol/L]
- Heel prick test in babies
What is phenylketonuria an example of?
A defect in amino acid metabolism (metabolism of phenylalanine in this case)
Homocystinuria
- Autosomal recessive
- Defect in cystathione B-synthase (CBS) enzyme OR defect in methionine synthase
- Excess homocysteine excreted in urine
- Excess homocysteine & methionine leading to symptoms
Symptoms of homocystinuria
- Dislocation of the lens
- Marfan-like stature (long limbs & fingers)
- Intellectual disability
Treatment of homocystinuria
- Low methionine diet (avoid nuts/peanut butter)
- Cysteine, Vit. B6, Betaine, B12 and Folate supplementation
- Avoid high protein food (milk, meat, fish, cheese, egg)
Diagnosis of homocystinuria
- Elevated plasma homocysteine + methionine
- Presence of oxidised homocysteine in urine
- Heel prick test in babies
In healthy adults, there is a steady state in which the amount of nitrogen taken into the body equals the amount of nitrogen lost. This is known as…
Zero nitrogen balance or N-balance
The nitrogen balance in periods of active growth, pregnancy, tissue growth, convalescence (muscle atrophy repair)
Positive N-balance (intake exceeds excretion)
The nitrogen balance in periods of starvation, malnutrition and trauma
Negative N-balance
Non-essential amino acids
Body can synthesise these
Most nitrogen leaves the body as…
- Urea (85%)
- Creatinine (5%)
- Ammonia (3%)
- Uric acid in urine (sweat & faeces)
- Direct loss of protein from body (skin, hair, nails)
Most nitrogen enters the body as…
Protein (>90%)
Where does the urea cycle take place?
Liver cytosol and mitochondria
Describe urea cycle steps in this diagram
Reaction 1 = carbamoyl phosphate is formed which is catalysed by carbamoyl synthetase enzyme.
Reaction 2 = carbamoyl portion of this product is transferred to ornithine .
Reaction 3 = citrulline is released to cytoplasm - combined with aspartate to form argininosuccinate.
Reaction 4 = argininosuccinate cleaved by lyase (ASL enzyme) to form arginine + fumarate.
Reaction 5 = arginine is cleaved by arginase to urea + ornithine.
Name the most common urea cycle disorder
Ornithine transcarbamylase deficiency (OCT)
Ornithine transcarbamylase deficiency (OTC)
- Example of urea cycle defect
- X-linked
Treatment of urea cycle defects (OTC)
- Low protein diet
- Replace amino acids in diet with keto acids
Symptoms of urea cycle defects (OTC)
Vomiting, lethargy, irritability, developmental delay, seizures, coma
Creatinine is a clinical marker of…
Renal function
Increased creatinine in the blood indicates…
Nephron (renal) damage
Which enzymes deaminate amino acids (get rid of excess ammonia)?
- Amino acid oxidases
- Glutaminase
- Glutamate dehydrogenase
Where does deamination occur?
Liver and kidney
Normal range of ammonia
25-40uM/L
In transamination - the NH2 from one amino acid is transferred to a keto-acid (a-ketogutarate) to produce a new amino acid (glutamate). This reaction is catalysed by…
Aminotransferases
In what disease processes may you expect to observe elevated plasma Alanine Aminotransferase (ALT) and Aspartate Aminotransferase (AST)?
Conditions that cause extensive cellular necrosis such as…
- Viral hepatitis
- Autoimmune liver disease
- Toxic injury
Alanine Aminotransferase (ALT)
Catalyses alanine -> glutamate [transamination]
Aspartate Aminotransferase (AST)
Catalyses aspartate -> glutamate [transamination]
Some amino acids cannot be synthesised in the body and are therefore essential and must be obtained within a diet. True or false?
True
The only two ketogenic amino acids are…
Lysine and leucine
A negative N-balance occurs in…
Burns, kwashiorkor, trauma, illness
Ammonia is ___ and it has to be converted to ___ and then excreted in the ___
Ammonia is toxic and it has to be converted to urea and then excreted in the urine
The urea cycle takes place in
Liver
What is the most common and safe form of ammonia which is transported from tissue via blood to the liver?
A) Glutamine
Ammonia converted to glutamine by glutamine synthase
The main site of urea synthesis is…
Liver
The most common defect in the urea cycle arises from the deficiency of the enzyme…
Ornithine transcarbamylase
Homocystinuria enzyme
Cystathione beta-synthase deficiency (CBS)
Methionine synthase
Phenylketonuria enzyme
Phenylalanine hydroxylase (PAH) deficiency
What enzyme deficiency is cause of most common urea cycle disorder (X-linked)?
Ornithine transcarbamylase (OTC) deficiency
Free amino acids in the body are used for…
A) Protein synthesis (~75%)
B) Synthesis of nitrogen-containing compounds (e.g., purines, creatine & haem)
C) Oxidation to energy
D) Synthesis of important signalling molecules from amino acids…
L-arginine → nitric oxide
L-cysteine → hydrogen sulphide
What hormone stimulates the breakdown of muscle protein in skeletal muscle (proteolysis)?
Cortisol
What amino acids are both glucogenic and ketogenic?
Isoleucine, threonine, phenylalanine, tyrosine and tryptophan
The two fates of N atoms in amino acids…
1) Transferred to other molecules (transamination)
2) Removed (deamination)
Free ammonia NH3 (NH4+) from the deamination of amino acids has two fates…
1) Excreted in the urine via urea cycle
2) Used to produce alanine and glutamine (ammonia detoxification)
Ammonia can be detoxified by converting the ammonium ion to ___ using the enzyme ___ ___
Ammonia can be detoxified by converting the ammonium ion to glutamine using the enzyme glutamine synthetase
Most N leaves the body as ___ (~85%), ___ (~5%), ammonia (~3%), uric acid (sweat, faeces),
What are the two fates of ammonia from the deamination of amino acids?
1) Excreted in the urine via urea cycle
2) Used to produce alanine and glutamine (ammonia detoxification)
How can ammonia be detoxified?
By converting the ammonium ion to glutamine using the enzyme glutamine synthetase.
What is the most common form of nitrogen leaving the body?
Most N leaves the body as urea (~85%), creatine (~5%), ammonia (~3%), uric acid (sweat, faeces), direct protein loss (hair, skin & nails).
How does most nitrogen enter the body?
As protein (>90%).
What is transamination?
A process involving the transfer of an amino group from one amino acid to a keto acid.
What is the clinical importance of transamination?
ALT and AST are important diagnostic markers
- ALT (Alanine Aminotransferase) and AST (Aspartate Aminotransferase) are key diagnostic markers for liver damage. They are enzymes that are found inside liver cells. When liver cells are damaged or inflamed, these enzymes leak into the bloodstream, leading to elevated levels of ALT and AST in blood tests.
What enzyme converts glutamate to alpha-ketoglutarate and ammonium ion (NH4+)?
Glutamate dehydrogenase.
What is the difference between a peptide/polypeptide and a protein?
Polypeptide = single linear chain of many amino acids held together by amide bonds.
Protein = generally >50 amino acids long.
What is the minimum categorization of proteins in terms of amino acid length?
Proteins can range from 40 to 100 amino acids long.
Where do carbon atoms for the synthesis of non-essential amino acids come from?
- Intermediates of glycolysis (C3)
- Pentose phosphate pathway (C4, C5)
- Krebs cycle (C4, C5)
What are the sources of the amino group for amino acid synthesis?
- Other amino acids by process of transamination
- From ammonia
What are conditionally essential amino acids?
Cysteine, Arginine, and Tyrosine are required by children and pregnant women due to high rates of protein synthesis.
What are major nitrogen-containing compounds?
- Proteins
- Amino acids
- Purines & Pyrimidines
What are minor nitrogen-containing compounds?
- Porphyrins (haem)
- Creatine
- Neurotransmitters (e.g., dopamine)
- Some hormones (e.g., adrenaline)
What is zero nitrogen balance?
Nitrogen intake = nitrogen output.
What is positive nitrogen balance?
Nitrogen intake > nitrogen output, indicating an increase in total body protein.
What is negative nitrogen balance?
Nitrogen intake < nitrogen output, indicating a net loss of body protein.
What are some causes of negative nitrogen balance?
Trauma, illness, burns, and malnutrition.
What is the breakdown product of creatine and creatine phosphate in the muscle?
Creatinine.
What are glucogenic amino acids?
All except leucine and lysine; can be converted into intermediates that feed into gluconeogenesis.
What are ketogenic amino acids?
Lysine and leucine.
Which amino acids are both glucogenic and ketogenic?
- Phenylalanine
- Tyrosine
- Tryptophan
- Threonine
- Isoleucine
What is a useful investigation that estimates muscle mass?
Creatinine urine excretion over 24h is proportional to muscle mass.
What is the largest store of fuel in a 70kg man?
Triacylglycerol (~15kg, 600,000kJ).
What is the smallest store of fuel in a 70kg man?
Glycogen (~0.4kg, ~4,000kJ).
How much does muscle protein make up in a 70kg man?
~6kg, ~100,000kJ.
What are the two main pathways by which nitrogen is removed from amino acids?
- Transamination
- Deamination
Why is the removal of nitrogen from amino acids essential?
- To allow the carbon skeleton of amino acids to be used in oxidative metabolism.
- Removed nitrogen can be incorporated into other compounds.
- Removed nitrogen is excreted from the body as urea.
What are aminotransferases?
Enzymes which catalyse transamination and are important diagnostic markers of liver function test (LFT).
What are two examples of amino acid metabolism disorders?
1) Phenylketonuria (PKU)
2) Homocystinuria.
What are the effects of defects in the urea cycle?
1) Hyperammonaemia
2) Accumulation/excretion of urea cycle intermediates.
