Amino Acids

Amino acids are not stored anywhere in the body.

Amino acids are obtained from diet, de novo synthesis, or produced from normal protein degradation. 

Protein turnover: the simultaneous synthesis & degradation of protein molecules. 

Its constant: rate of syn = rate of degradation.

AKA nitrogen balance!

KO mouse: genetically engineered mouse in which researchers have inactivated an existing gene and replaced/disrupted it with artificial DNA.

Purpose: Important for studying functions of genes that have been sequenced.

• 1. Short-lived proteins; minutes/hours
• 2. Long-lived proteins; days to weeks 
• 3. Structural proteins; months to years. 

• 1. Target specific gene to eliminate exons & insert Neo gene for selection
• 2. Without exons, gene codes for defective protein.
• 3. Select embryonic stem cells that contain the targeted defective gene by growing with antibiotic. Cells WITH Neo (neomycin) will grow. Cells w/o will die. 
• 4. Inject ES cell into blastocyst
• 5. Implant blastocyst into pseudo-pregnant mouse
• 6. Mouse will have some tissues derived from targed ES cells. Need targeted gonads to mouse will give birth to KO pup.

• 1. Proteins
• 2. Urea (excreted in urine)
• 3. Ammonia - used in synthesis of AA, nucleotides, biological amines, or excreted in urine.
• 4. A-keto acids - intermediates in metabolism. 

From: dietary protein, de novo synthesis of nonessential protein, and protein breakdown from body.

Goes to: synthesis of body protein, amino acids consumed as precursors of essential N-containing molecules and conversion of AA to ketogenic, gluconeogenic (glucose, glycogen, FAs, ketone bodies, or CO2+H2O). 

In cells, blood and EC fluids

• 1. Glucose
• 2. Glycogen
• 3. FAs
• 4. Ketone bodies
• 5. Doesn't really count, but CO2 + H2O.

ALT = alanine amino transferase

Converts pyruvate into alanine (amino acid) or vice versa (gluconeogenic), while converting glutamate to a-ketoglutarate.

• 1. Pyruvate
• 2. Ketogenesis (lysine & leucine only). Produce acetyl CoA or acetoacetyl CoA CANNOT TURN INTO GLUCOSE. 
• 3. Gluconeogenesis
• 4. Oxidation - during starvation, the reduced C skeleton is oxidized to CO2 and H2O thereby producing ATP.

During starvation, the reduced C skeleton is oxidized to CO2 and H2O thereby producing ATP.

Isoleucine, phenylalanine, threonine, tryptophan, and tyrosine. 

IPTTT

Lysine & leucine

Produces acetyl CoA or acetoacetyl CoA

NO, they are solely ketogenic. 

• 1. Cannot be synthesized by humans at all
• 2. Cannot be synthesized in sufficient amounts by humans. 

Arginine (urea synthesis), methionine (cysteine), and phenylalanine (tyrosine). 

Tyrosine: If not enough phenylalanine is consumed

Cysteine: if not enough methionine is consumed. 

PKU --> cannot process phenylalanine, so put on a phenylalanine restricted diet, so tyrosine becomes essential. 

400 g turned over daily

20 g of protein --> digestive enzymes or lost from sloughing of gut epithelial cells. 

Steady state: rate of protein syn = rate of protein degradation

Positive N balance: rate of protein syn>rate of protein degrad (growth, pregnancy, and recovery from illness). 

Negative N balance: rate of protein syn < degradation (illness, starvation). 

Proteins are hydrolyzed to AA by pepsin (stomach protease) & pancreatic enzymes (chymotrypsin & trypsin), 

Absorption: AA are absorbed into hepatic portal blood, taken up by liver, and either used or shuffled elsewhere.

1. Lysosomes (ATP indep degradative enzyme systen of the lysosomes)  - general housekeeping, routine degradation of proteins from EC cells via phagocytosis & cell receptors

2. Autophagy - last resort (prevents cell death by recycling noncritical proteins & uses AA to synthesize more important proteins).

3. Ubiquitination - tags proteins for degradation (ATP-dep) in cytosol. Ubiquitin-proteasome system. 

• 1. Ubiquitination: Proteins to be degraded are first covalently attached to ubiquitin.
• 2. Ubiquitinated protiens are recognized by cytosolic proteasome
• 3. Proteasome unfolds, deubiquitinates and transports protein to its proteolytic core (ATP dep). 
• 4. Peptide fragments produced by proteasome are degraded to AA in the cytosol. 

Ubiquitin is recycled. The last enzyme: non-specific proteases that split peptide fragments into amino acids. 

Proteasomes degrade endogenous proteins

Lysosomal enzymes (acid hydrolases) degrade EC proteins

ex. plasma proteins taken into cell by endocytosis & cell-surface membrane proteins that are used in receptor-mediated endocytosis. 

In healthy cells, lysosomes are non-selective.

During prolonged fasting, a special pathway is activated that selectively degrades cytosolic proteins containing a Lys-Phe-Glu-Arg-Gln pentapeptide sequence. 

LPGAG

Function: (1) degradation of cellular proteins/organelles (2) to recycle nutrients (3) or break down damaged/toxic materials. 

Vacuole forms around cytoplasmic constituents and fuses with lysosomes where contents are degraded by vacuolar hydrolases and proteases. 

The N-terminal amino acid. 

yes. 

Autophagy increases.

Increases breakdown of nonvital proteins, releasing nutrients, ensuring vital processes can continue.

Rapamycin suppresses immune system and is valuable for transplant surgeries by acting on mTOR (mammal target of rapamycin). 

Found on Easter Island

Rapamycin activates autophagy and inhibits mTOR.

mTOR (mammalian target of rapamycin) cell growth regulator that integrates growth factor & nutrient signals. 

• Activated by insulin & AA --> promotes growth
• Inhibited by rapamycin and nutritiond deficit.

• 1. Induction - external stimuli (starvation, nutrient depletion, ischemia, rapamycin) inhibits mTOR.
• 2. Autophagosome formation - cytosolic proteins/organelles are sequestered by a 2xmembrane vesicle (maybe from ER?)
• 3. Docking and fusion with the lysosome
• 4. Breakdown  of autophagic vesicle and relase of contents (AA, nutrinets, etc) for recycling in cell.

Autophagy is activated by rapamycin and starvation/nutrient depletion

Inhibited by mTOR (which is activated by insulin/AA). 

Found in nucleus & cytoplasm

Degrade unneeded/damaged proteins and respond to cellular stresses (infection or oxidative damage). 

Ubiquitin-proteasome degradation process

• 1. Attachment of additional ubiquitin molecules
• 2. N-terminal residue (met/pro = slow; arg/leu = fast)
• 3. Presence of PEST sequences (Pro-Glu-Ser-Thr) speed up process. 

ubiquitin-proteasome degradation.

lysosomal does NOT require ATP. 

• 1. Digestion of autophagic vesicles
• 2. Phagocytosis (ingestion of other dying cells/EC material)
• 3. Endocytosis (recycling of cell-surface receptors, LDL)

• 1. Body protein (400 g/day)
• 2. Glucose
• 3. CO2+ATP
• 4. Ketone bodies/FAs/steroids
• 5. Synthesis of purines, pyrimidines, NTs, porphyrins, creatinine
• 6. Urea cycle

Glutamine & alanine are carriers of NH3.

From muscle/tissues to liver. 

TRICK QUESTION. Mostly liver, but branched amino acids (isoleucine, leucine, valine) are preferentially metabolized in MUSCLE.

AA undergoes degradation when AA intake exceeds biosynthetic needs. AA undergoes GNG when protein is needed as a fuel source.

Goal: to separate a-amino group from C skeleton and shunt each into specialized pathways. 

Amino group --> liver --> urea; C skeleton --> pyruvate, acetyl CoA or TCA intermediates. 

B6 (pyridoxal phosphate) as intermediate & transaminase

A-ketoglutarate, except lysine & threonine. 

They help facilitate creation of energy, but they play an indirect role, so in themselves, do not create energy.

AST: Aspartate aminotransferase; OAA; transfers amino group from glutamate to OAA forming aspartate for urea cycle. 

Yes, this is reversible. 

High levels signify damage to cells rich in these enzymes (liver, muscle, etc). 


Could indicate liver disease, myocardial infarction, hepatitis, toxic injury, etc. 

ALT is more specific than AST for liver disease, but AST is more sensitive bc liver has high levels of AST. 

• 1. Help maintain adequate levels of non-essential AA required for protein synthesis
• 2. Funnel AA groups from catabolized AA to glutamate & aspartate for eventual excretion as urea. 

Needed for transamination for syn of nonessential AAs, production of glucose, and loss of amino group to urea. 

+ N Balance and - N balance both increase transamination because these reaction are all reversible. 

Bc BCAA are broken down in muscle and high BCAA in blood may directly promote insulin resistance.