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Guide e consigli
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Proteins' Metabolism, Appunti di Biochimica Metabolica

- un'introduzione sugli amino acidi - digestione delle proteine parlando degli enzimi che aiutano il processo. - "Endopeptidases" e "Exopeptidases" sono spiegati in modo chiaro specificando i loro meccanismi e il loro ruolo nei diversi organi del corpo durante la digestione. - Amino Acid Transport - "Clinical Relevance" e dei problemi che si potrebbero avere se qualche Aminoacido non fosse presente. - catabolismo delle proteine e dei processi metabolici che avvengono, distinguendo di parte in parte (liver, skeletal muscle, kidneys...) - "protein turnover" e affrontare il discorso di "ubiquitin". processi di "Transamination" e "Oxidative Deamination" e degli enzimi più importanti. - ruolo dell'Ammonia (e dell'Urea Cycle) , della Glutamine Synthetase Pathway, del Glucose Alanine Cycle. - Heme Catabolism, Catecholamines, Histamines, Serotonin, Creatine, Melatonine, Nitric Oxide Materia: Biochemistry, primo anno di medicina, professore Luigi Anastasia UniSr

Tipologia: Appunti

2023/2024

In vendita dal 11/10/2024

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ANASTASIA PROTEINS
Proteins are involved in nearly every physiological process in the body. They function in
various capacities, including:
Transport Proteins: e.g. hemoglobinbin (transports oxygen), ferritin (transports iron)
Structural Proteins: e.g., elastin and collagen (support the body)
Contractile Proteins: e.g., actin and myosin (required for movement)
Enzymes: Catalyze numerous biochemical reactions
Hormones: Regulate various bodily functions
Receptors: e.g., proteins involved in nerve cell transmission
Transcription Factors: Regulate gene expression
These functions highlight the critical roles of dietary proteins in maintaining human health
and physiological processes.
AMINO ACIDS IN PROTEINS
- Number of amino acids: 20 standard aminos
- Structure: carbon atom linked to a hydrogen atom, amine group, carboxyl group and
an alkyl side chain
We need many enzymes to process these proteins in different ways.
NUTRITIONAL CLASSIFICATION
- Essential amino acids
- Non essential amino acids
- Conditionallly essential amino acids
Digestion:
If we break down these peptide bonds
we get shorter chains, during the
digestion we have a series of proteases
that we call peptidases. These
peptidases will catalyze the hydrolysis
of the peptide bond.
When we think about the digestive
system we start from the mouth…
In the mouth not a real digestion occurs
, we just have a mechanical process in
which with the chewing we are making
the food easier to be attacked later.
We are dropping the pH at about 2 or 3
(important) so we can kill the bacteria in
our food, and proteins will start to
denature and the peptide bond will
become more accessible to the
enzymes.
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ANASTASIA PROTEINS

Proteins are involved in nearly every physiological process in the body. They function in various capacities, including: ○ Transport Proteins: e.g. hemoglobinbin (transports oxygen), ferritin (transports iron) ○ Structural Proteins: e.g., elastin and collagen (support the body) ○ Contractile Proteins: e.g., actin and myosin (required for movement) ○ Enzymes: Catalyze numerous biochemical reactions ○ Hormones: Regulate various bodily functions ○ Receptors: e.g., proteins involved in nerve cell transmission ○ Transcription Factors: Regulate gene expression These functions highlight the critical roles of dietary proteins in maintaining human health and physiological processes. AMINO ACIDS IN PROTEINS

  • Number of amino acids: 20 standard aminos
  • Structure: carbon atom linked to a hydrogen atom, amine group, carboxyl group and an alkyl side chain We need many enzymes to process these proteins in different ways. NUTRITIONAL CLASSIFICATION
  • Essential amino acids
  • Non essential amino acids
  • Conditionallly essential amino acids Digestion: If we break down these peptide bonds we get shorter chains, during the digestion we have a series of proteases that we call peptidases. These peptidases will catalyze the hydrolysis of the peptide bond. When we think about the digestive system we start from the mouth… In the mouth not a real digestion occurs , we just have a mechanical process in which with the chewing we are making the food easier to be attacked later. We are dropping the pH at about 2 or 3 (important) so we can kill the bacteria in our food, and proteins will start to denature and the peptide bond will become more accessible to the enzymes.

DIGESTION AND ABSORPTION OF PROTEIN: MOUTH

Unless you are eating it raw, the first step in digesting an egg (or any other solid food) is chewing. The teeth begin the mechanical breakdown of large egg pieces into smaller pieces that can be swallowed. The salivary glands secrete saliva to aid swallowing and the passage of the partially mashed egg through the esophagus. No protein digestion occurs in the oral cavity. The first phase of protein digestion takes place in the stomach through the action of an endopeptidase, pepsin. Pepsin is a hydrolytic enzyme that attacks the peptide bonds in dietary proteins, specifically those formed by aromatic amino acids such as tyrosine and phenylalanine. We are not really able to store proteins or amino acids! We need to use them for biosynthesis or to convert them into carbon chains that do not contain hydrogen. There are different types of enzymes!! Whole proteins are not absorbed –> they are too large to pass through cell membranes intact. DIgetìstive enzymes. Hydrolases break peptide bonds. They are secreted as inactive pre-enzymes and prevent self digestion. These are all the enzymes that we could encounter. This digestive process requires several steps. ENDO VS EXO PEPTIDASES the key difference is that the endopeptidase breaks peptide bonds within the protein molecules while the exopeptidase cleaves peptide bonds at the terminals of the protein molecules. They start to release progressively the amino acids from the end of the peptide chain and allow to get to the free amino acids. Some are at the N terminus and some are at the C terminus, they will act on different types and shapes of bonds. They need to be secreted in an inactive form as we said.

PEPSINOGEN AND PEPSIN

Pepsin is produced in an inactive form known as pepsinogen by the chief cells of the gastric mucosa. The release of pepsinogen is stimulated by the hormone gastrin in response to HCl. The activation of pepsinogen into pepsin occurs due to the action of HCl in the gastric environment and autocatalytically. This activation involves the removal of a peptide segment from pepsinogen. The low pH will cut the pepsinogen which is longer than pepsin, and pepsin will be activated with the hydrolysis of pepsinogen. A KEY ROLE is played with HCl. How do we synthesize it? https://youtu.be/opWBDkgiWx4?si=d14MmBykgWDl9JEa

MUCUS LAYER IN THE STOMACH

The stomach produces a layer of mucus gel rich in bicarbonate, which protects it

from self-digestion. The mucus is made of mucins, glycoproteins rich in

carbohydrates, and also contains phospholipids. The mucus covers and protects the

mucosal surface of the stomach and duodenum.

CLINICAL RELEVANCE - PEPTIC ULCER DISEASE

If for some reason the mucus layer is breached, the epithelial cells are exposed to

concentrated stomach acid and the digestive enzymes contained within the gastric

juices. As the stomach wall is made up of the same proteins and lipids as many

foods we eat, if this mucosal barrier is broken the stomach will begin to digest itself,

forming a peptic ulcer.

There are certain precipitations to this condition, including gastric colonization of

Helicobacter pylori, exposure to nonsteroidal anti-inflammatory drugs (NSAIDs) such

as ibuprofen, and excess acid secretion as seen in Zollinger-Ellison syndrome.

DIGESTION OF PROTEINS IN THE INTESTINE BY PANCREATIC ENZYMES

→ HORMONAL REGULATION

as acidic stomach contents pass into the DUODENUM, the low pH triggers the

secretion of hormones:

- Secretin: stimulates the pancreas to secrete bicarbonate

- Cholecystokinin (CCK): stimulates the secretion of pancreatic enzymes

Pancreatic Enzymes:

- Endopeptidases: trypsin, chymotrypsin, elastase

- Exopeptidases: carboxypeptidases, aminopeptidases

MECHANISM OF ENDOPEPTIDASES

SERINE PROTEASES:have a serine in their active site that covalently attaches to

one of the protein fragments as an enzymatic intermediate. This class includes the

chymotrypsin family (chymotrypsin, trypsin, and elastase), and the subtilisin family.

CYSTEINE PROTEASES: have a similar mechanism, but use cysteine rather than

serine. These include the plant proteases (papain, from papaya, and bromelain, from

pineapple) as well as mammalian proteases such as calpains.

ASPARTIC PROTEASES: have two essential aspartic acid residues that are close

together in the active site, although far apart in the protease sequence. This family

includes the digestive enzymes pepsin and chymosin.

METALLOPROTEASES: use metal ion cofactors to facilitate protein digestion and

include thermolysin. Finally, the fifth class of proteases, threonine proteases, has an

active-site threonine.

TRYPSIN

is secreted in the zymogen form (tripsinogen) and activated by enterokinase and

trypsin itself. Is specific for cleaving

peptide bonds contributed by lysine,

arginine (basic aminos) and acts on the

zymogen forms of other pancreatic

enzymes and activates them. It has a

weak action on casein.

CHYMOTRYPSIN AND ELASTASE

CARBOXYPEPTIDASE

EPITHELIAL CELLS PEPTIDASES

Epithelial Cell Activity Brush-Border Peptidases:

  • Located on the surface of intestinal epithelial cells.

•Amino acids cannot be stored but are used for protein synthesis. •Excess amino acids are broken down to produce energy. •After absorption by intestinal cells, amino acids enter the bloodstream (via the portal vein) and are distributed to all cells in the body. IMPORTANT!! not all amino acids are the same, they have different side chains and different structures. They can be essential or non essential. The categorisation of amino acids into their essential, conditional and non-essential groups in humans Conditional amino acids are not usually essential amino acids, only in times of illness and stress. Essential amino acids are not produced naturally by the body and must come from dietary intake whereas non-essential amino acids are produced by our body WHY CAN’T WE MAKE SOME? Humans have lost the ability to synthesize nine essential amino acids due to genetic mutations in our ancestors. These mutations affected enzymes necessary for complex synthesis pathways. The missing enzymes impact the conversion processes shown in the yellow, green, red, and blue boxes on the chart.

  • Yellow Box (Bottom Left): Pyruvate to leucine and valine.
  • Green Box (Left): Aspartate to multiple amino acids.
    • Red Box (Right): Glutamine to histidine.
    • Blue Box (Top Right): Synthesis of tryptophan and phenylalanine. Due to these mutations, humans must obtain these essential amino acids from their diet.

WHAT HAPPENS WHEN ESSENTIAL AMINOS ARE MISSING?

  1. Protein Breakdown: The body is forced to break down its own proteins to obtain essential amino acids.
  2. Effects of Prolonged Fasting During extended periods of fasting:
  • The liver can lose up to 50% of its proteins.
  • Skeletal muscle can lose up to 30% of its proteins.
  • The heart loses only about 3% of its proteins.
  1. Amino Acid Reserve:
  • The liver and skeletal muscles act as a reserve of amino acids, providing them to the body when needed. (ESSENTIAL) AMINOS DEFICIENCY Symptoms of Lacking Amino Acids ● Problems with Thyroid: Impaired thyroid function leading to hormonal imbalances. ● Drop in Performance: Reduced physical performance and endurance. ● Being Tired: Chronic fatigue and low energy levels. ● Weakness and Pain in Muscles: Muscle weakness and soreness, difficulty in muscle recovery. ● Losing Hair: Hair loss and thinning BRANCHED CHAIN AMINO ACIDS (BCAAs) ● Branched-Chain Amino Acids (BCAAs): Also known as branched-chain amino acids. Include leucine, isoleucine, and valine. Importance ● Essential for Humans: • Represent about 35% of essential amino acids in muscle proteins. • Account for 40% of the required amino acids in mammals. Functions • Energy Production: Can be broken down (catabolized) to produce energy (gluconeogenesis). Important for endurance sports. ● Protein Synthesis: Play a crucial role in protein synthesis. Beneficial for strength sports and bodybuilding to increase muscle mass. AMINO ACID CATABOLISM Schematic diagram of the metabolism of amino acids, including the 3 major pathways: reutilization in the synthesis of new proteins, union with cofactors to produce amino acid derivatives, and catabolism. Catabolism of amino acids includes

NH4+ is either excreted or used in the urea cycle for arginine synthesis.

  • Acid-Base Balance Acts as an important buffer against acidosis PROTEIN TURNOVER Definition Protein Turnover: The process of degradation and resynthesis of proteins. Half-Lives of Proteins
  • Varies Widely: Can range from several minutes to many years. Types of Proteins
  • Structural Proteins:
  • Usually stable.
  • Example: Lens protein crystallin, which lasts the whole life of the organism.
  • Regulatory Proteins:
  • Short-lived.
  • Their rapid turnover allows quick changes in metabolic processes. HOW CAN WE DISTINGUISH PROTEINS THAT ARE MEANT FOR DEGRADATION Ubiquitin Tagging → Ubiquitin: A small (8.5-kd) protein found in all eukaryotic cells. Acts as a tag marking proteins for destruction. Known as the "black spot," signaling proteins for degradation. Structure of Ubiquitin Extended Carboxyl Terminus: Glycine residue that links ubiquitin to target proteins.
  • Lysine Residues: Allow the formation of polyubiquitin chains by linking additional ubiquitin molecules. DECARBOXYLATION OF AMINO ACIDS → transform carboxylic group into CO Just remember that one way to synthesize amines is starting from the amino acid and decarboxylation. It is catalyzed by decarboxylase enzymes and they require PLP as a coenzyme. We can find them in various tissues. FATE OF ALPHA AMINO GROUP OF AMINO ACIDS
  • Protection from Oxidative Breakdown: The presence of the α-amino group keeps amino acids safely locked away from oxidative breakdown.
  • Obligatory Step in Catabolism: Removal of the α-amino group is an essential step in the catabolism of all amino acids. •Nitrogen Removal: Once removed, this nitrogen can be incorporated into other compounds or excreted. The remaining carbon skeleton is metabolized.
  • Excretion of Excess Nitrogen: Different animals excrete excess nitrogen in different forms: Ammonia, Uric Acid, Urea DETACHMENT OF THE AMINO GROUP The detachment of the amino group can occur through: 1.Transamination:
    • Process: Transfer of an amino group from an amino acid to an α-keto acid.
  • Example: Alanine + α-Ketoglutarate → Glutamate + Pyruvate 2.Oxidative Deamination:
  • Process: Removal of an amino group as ammonia (NH4+) and conversion to a keto acid.
    • Example: Glutamate → α-Ketoglutarate + NH4+ + NAD(P)H/H+ SIGNIFICANCE OF TRANSAMINATION Dual Purpose of Transamination 1.Interconversion of Amino Acids: Allows the conversion of one amino acid into another. 2.Energy Utilization: Directs excess amino acids towards energy production. NOT GOING TO ASK WHICH AMINO ACIDS CAN BE CONVERTED INTO WHICH DERIVATIVES (TRANSAMINATION ALREADY DONE ON MY PAPER SHEETS) TRANSAMINATION MECHANISM
  • An amino group (NH3+) from an amino acid is transferred to an α-keto acid
  • This produces a new amino acid and a new α-keto acid. The enzymes involved are TRANSAMINASES or AMINOTRANSFERASES
    • PLP is used as coenzyme, which is the active form of vitamin B PLP acts as a carrier of amino groups at the active site of aminotransferases. TRANSAMINATION: ALT and AST They are very found in the liver and in the skeletal muscle. The key enzymes are:
    • Aminotransferase Catalyzes the transamination reaction, requires pyridoxal Phosphate PLP as a cofactor Example Reactions:
    • ALT (alanine aminotransferase) or GTP

OXIDATIVE DEAMINATION (ALREADY DONE ON MY PAPERS)

STIMULI:

D-Amino acid oxidase

● Removal of the α-amino group is the first step in catabolism of amino acids. It

may be accomplished oxidatively or nonoxidatively.

● Oxidative deamination is stereospecific and is catalyzed by L- or D-amino acid

oxidase.

● The initial step is removal of two hydrogen atoms by the flavin coenzyme, with

formation of an unstable α-amino acid intermediate.

● This intermediate undergoes decomposition by addition of water and forms

ammonium ion and the corresponding α-keto acid: L-amino acid oxidase

occurs in the liver and kidney only

SUMMARY TRANSAMINATION AND OXIDATIVE DEAMINATION

AMMONIA TRANSPORT TO THE LIVER

there are two primary mechanisms by which ammonia is transported to the liver for detoxification and excretion

  1. GLUTAMINE SYNTHETASE PATHWAY
  • Muscle: ammonia is combines with glutamate to form glutamine by the enzyme GLUTAMINE SYNTHETASE. This reaction REQUIRES ATP
  • Liver: glutamine is transported to the liver, where the enzyme GLUTAMINASE onverts it back into glutamate and free ammonia. GDH then converts glutamate into alphaKG into the urea cycle to form
  • Urea formation: the ammonia is then incorporated into the urea cycle to form urea, which is excreted by the kidneys
  1. GLUCOSE ALANINE CYCLE
  • Muscle: pyruvate transaminates with glutamate to form alanine and alphaKG .This reaction is catalyzed by ALT
  • Liver: alanine is transported to the liver, where ALR reverses the reaction to form pyruvate and glutamate. GDH then converts glutamate to alphaKG and ammonia
  • Urea formation: as in the glutamine pathway, the ammonia enters the urea cycle to be converted into urea for excretion

AMINO ACIDS, AMMONIA AND HEPATIC ENCEPHALOPATHY

A schematic summary of pathophysiology of

hepatic encephalopathy and

ammoniogenesis in end-stage liver disease

patients.

As a result of liver failure and cirrhosis, less

ammonia is detoxified in the

liver and portosystemic shunting allows a

greater amount of ammonia to enter the

systemic circulation bypassing the portal

system.

In cirrhotic patients, ammonia detoxification

takes place less in the liver

and more in the skeletal muscle and kidney.

In liver, ammonia is converted

to urea and then excreted through the

kidneys and into the colon.

In skeletal muscle and kidney, ammonia is

converted to glutamine thus

enzymatically removing the ammonia.

In addition, astrocytes in the brain also play a role in detoxifying ammonia

via glutamine synthetase, the increase in ammonia concentrations passing

the blood–brain barrier results in increase in glutamine in the brain and this

bring in more water into astrocytes.

This swelling of astrocytes leads to cerebral edema and intracranial

hypertension, ultimately resulting in neuronal dysfunction

ROLE OF GABA IN HEPATIC ENCEPHALOPATHY

1. NEURONAL IMPACT:

Elevated ammonia increases glutamate formation, leading to more GABA production in neurons. GABA is an inhibitory neurotransmitter that calms neuronal activity

  1. HEPATIC ENCEPHALOPATHY Elevated GABAergic activity contributes to the neurological symptoms of hepatic encephalopathy. Symptoms include confusion, altered level of consciousness and coma in severe cases SKELETAL MUSCLE, KIDNEY AND LIVER INTERPLAY Muscle
  • protein breakdown
  • glutamine and alanine production Liver
  • alanine metabolism
  • rea Cycle

Kidney

  • Ammonia Excretion
  • Alanine transport Gut
  • Glutamine utilization

KIDNEY AND GLUTAMINE

● While the main function of the kidney is the regulation of blood solutes, it also

serves to replenish glucose stores through de novo gluconeogenesis using

amino acids as precursors and maintains acid-base balance

● Glutamine is an important substrate for both functions, with its catabolism

generating the energy intermediate α-ketoglutarate and the weak base

bicarbonate.

● Its importance is highlighted by the fact that renal glutamine uptake accounts

for as much as 58% of total renal amino acid uptake and assessments of

renal arteriovenous differences consistently demonstrate a large absorption of

glutamine

GLUCOSE ALANINE CYCLE

Muscle:

- Glycolysis

- Transamination

- Transport

Liver

- Transamination

- Gluconeogenesis

- Urea Cycle

LIVER AND AMMONIA

METABOLISM OF AMINO ACIDS IN

THE LIVER

● The liver is the main site of amino

acid catabolism. Amino acid

absorbed from the gut are carried

to the liver via the portal vein.

● Most amino acids are deaminated

via transamination to generate glutamate and the corresponding α-ketoacid of

the original amino acid.

● Depending on whether the amino acid is

glucogenic or ketogenic, the α-ketoacid

will enter gluconeogenic or TCA cycle

pathways.

● Glutamate is then deaminated to

generate a free molecule of ammonia

and α-KG.

diffusion, with the rate of absorption depending on urine flow rate and the

degree of hydration.

● Additionally, 10% of urea is excreted through the gastrointestinal (GI) tract

and skin.

● Blood urea levels depend on several factors, including renal function and

perfusion, the protein content of the diet, and the amount of protein

catabolism.

UREA CYCLE DISEASES

The symptoms of different urea cycle disorders can overlap because they similarly

affect the body, making the diagnosis methods for these disorders also similar. The

illustrated urea cycle figure shows enzymes (in blue) and transporters (in red)

converting ammonia nitrogen into urea. Nitrogen from ammonia and aspartate is

transferred through several intermediate compounds (in green) before becoming

urea. A deficiency in any urea cycle enzymes or transporters results in the body's

inability or reduced ability to dispose of nitrogen, leading to ammonia accumulation.

OTHER USES OF AMINO ACIDS

1. PORPHYRIN METABOLISM

The most common metalloporphyrin is HEME, an HEME group consists of an iron Fe

ion held in a heterocyclic ring.

Hemoglobin synthesis is needed and is erythropoietic, Cytochrome synthesis is

needed too and is hepatic

Heme synthesis occurs in all cells due to the requirement for heme as a prosthetic

group on enzymes and electron transport chain proteins. By weight, the major

locations of heme synthesis are the liver (cytochrome p450) and the erythroid

progenitor cells (Hemoglobin) of the bone marrow.

Through a series of enzymatic steps involving URO I synthase, URO III co-synthase,

and URO decarboxylase, porphobilinogen is converted into uroporphyrinogen III,

which then progresses to coproporphyrinogen III

Lead can inhibit Ferrochelatase, disrupting the final step of heme synthesis. This

highlights the importance of each enzyme in the pathway and the potential

consequences of their inhibition.

HEME BIOSYNTHESIS DISORDERS: INHERITED AND ACQUIRED PORPHYRIA

→ affected individuals accumulate porphyrins in RBC, body fluids, liver and in certain

case in the brain. According to the type of porphyria, intermittent neurological

impairment, abdominal pain, hypertrichosis

HEME CATABOLISM

2. CATECHOLAMINES

They are synthesized from Tyrosine and stored in secretory vesicles.

Tyrosine is hydroxylated by tyrosine hydroxylase (rate limiting step) to form DOPA,

which is then decarboxylated by DOPA decarboxylase to form DOPAMINE which is

then hydroxylated by dopamine beta-hydroxylase to give NOREPINEPHRINE.

EPINEPHRINE is formed by N-methylation reaction using S-adenosylmethionine as

a methyl donor.

!!Parkinson’s disease is caused due to the production of insufficient dopamine

synthesis in the brain.

!!NADPH is the electron donor for the two hydroxylation steps