Biochemistry Flashcards
Amino acids - essential (obtained from dietary)
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TRYptophan
THreonine
HIStidine (basic)
Valine (branched)
Isoleucine (branched)
Phenylalanine
Methionine
(a)
Leucine (ketogenic) (branched)
Lysine (ketogenic) (basic)
Amino acid side chains - properties
Amino acids are organic compounds composed of an amine group (H3N+ - becomes ammonia if free), Carboxylic acid (CO2), Unique side chain (finger print) and
What are transamination reactions?
REVERSIBLE REACTIONS by which cells transfer the amino group from an amine or α-a.a. to an α-keto carboxylic acid. Enzymes involved in this reaction are transaminases/ ainotransferases. REQUIRES PYRIDOXINE (Vitamin B6) as a co-factor.
Cori cycle / Lactic acid cycle
Lactate and glucose exchange between muscles and the liver. (As high levels of lactate can lead to acidosis)
Involved metabolic pathways: glycolysis, anaerobic metabolism with lactate production, gluconeogenesis
Types of lactate dehydrogenase (LDH)
LDH 1, LDH 2 : Cardiac muscle, Erythrocytes, Kidneys (raised in MI, myocarditis, hemolysis, renal infarction - can be used in delayed dx MI instead of trop as lasts longer in blood)
LDH 3: Lungs, Lymphatic system, Platelets (raised in PE or platelet destruction)
LDH 4, LDH 5: Liver, Skeletal muscles (raised in liver disease, skeletal muscle injury, malignancy)
Poor specificity
The LDH enzyme exists in five isoenzymes (LDH1–LDH5) distributed across different tissues.
The direction of lactate ↔ pyruvate conversion depends on the cellular ratio of NAD+ to NADH.
Lactate dehydrogenase reaction
Pyruvate >< Lactate
NAD+ excess: Lactate > Pyruvate
NADH + H+ excess: Pyruvate > Lactate
Glycolysis - Reactions
Occurs int he cytosol of cells
Glucose (C6) to Pyruvate (2 x C3) through 10 steps. Pyruvate then converted to Acetyl-coA and can enter the citric acid cycle.
- Glucose Phosphorylation: Glucose → Glucose-6-phosphate (via hexokinase, irreversible, 1 x ATP used).
- Isomerization: Glucose-6-phosphate → Fructose-6-phosphate (via glucose 6 phosphate isomerase).
- Second Phosphorylation: Fructose-6-phosphate → Fructose-1,6-bisphosphate (via phosphofructokinase-1, irreversible, rate limiting step, 1 x ATP used; total EB -2ATP).
- Cleavage: Fructose-1,6-bisphosphate → isomers: Glyceraldehyde-3-phosphate (GAP) + Dihydroxyacetone phosphate (DHAP) (via aldolase).
- Isomerization: DHAP ↔ GAP (via triose phosphate isomerase). Only GAP can be further metabolised in glycolysis, resulting in 2 x GAP.
- Oxidation and phosphorylation: 2 x GAP → 2 x 1,3-Bisphosphoglycerate (via GAP dehydrogenase, NAD⁺ reduced to NADH plus H+; total EB -2ATP and 2(NADH plus H+)).
- ATP Formation: 1,3-Bisphosphoglycerate → 3-Phosphoglycerate (via phosphoglycerate kinase, ATP produced).
Substrate-level phosphorylation CREATES ENERGY (+2ATP; +2 (NADH and H+). - Phosphate Shift: 3-Phosphoglycerate → 2-Phosphoglycerate (via phosphoglycerate mutase).
- Dehydration: 2-Phosphoglycerate → Phosphoenolpyruvate (via enolase, water released).
- ATP Formation: Phosphoenolpyruvate → Pyruvate (via pyruvate kinase, irreversible ATP produced), irreversible
Net ATP gain = 2 ATP (4 produced, 2 used).
NADH plus H+ produced = 2 molecules per glucose.
https://www.youtube.com/watch?v=9HqCTiGzqfU&list=PLkv9qVBSWseGHDrmzrPmeaQC3bKxIPeUh&index=2
What is Substrate-level phosphorylation ?
When a phosphoryl group is transferred from a substrate to ADP or GDP to form ATP or GTP, coupled with the release of free energy.
Glycolysis stage 7
What happens to pyruvate after glycolysis
Pyruvate → acetyl-CoA (via pyruvate dehydrogenase) which is then
(a) passed on to citric acid cycle
(b) used as building block in other metabolic processes e.g. FA synthesis
Glycolysis - 3 regulatory enzymes
Hexokinase (Step 1)
Phosphofructokinase-1 (Step 3)
Pyruvate Kinase (Step 10)
Glycolysis - Regulations
Regulatory mechanisms
(1) Hormonal: via signalling molecules e.g. cAMP
(2) Allosteric: via regulatory molecules e.g. reactants or products
Hexokinase (Step 1)
- Glucagon inhibits transcription (increasing blood glucose levels, reducing cell glucose levels)
- Insulin facilitates activity of glucose transporters (more glucose in cells) and transcription of hex (activates first step of glycolysis)
- G6P inhibits hexokinase- neg feedback prevents excessive phosphorylation of glucose when G6P accumulates, avoiding unnecessary ATP consumption. G6P does not inhibit glucokinase (AKA hexokinase-IV liver-specific isoform), which allows the liver to continue processing glucose for glycogen synthesis or other pathways despite G6P accumulation.
- Glucose affinity of hexokinase changes in tissue. Glucokinase / Hexokinase - IV LOW AFFINITY, Hexokinase (extra-hepatic) HIGH AFFINITY - so when low levels of glucose, does not go to the liver.
Phosphofructokinase-1 (Step 3) RATE LIMITING STEP.
Inhibited by 3 factors:
(a) ATP (when energy cellular supply is sufficient)
(b) Citrate (accumulates in 1st step citric acid cycle when excess acetyl coA)
(c) Protons (acidic pH)
Activated by 2 factors:
(a) ANP (signifies a metabolic state in which glycolysis needs to be activated), counteracts ATP
(b) Fructose 2, 6-bisphosphate (completely different from fructose 1, 6-bisphosphate) - hormonal reg via cAMP which makes sure e.g. heart increase, whereas liver reduce.
Pyruvate Kinase (Step 10)
Inhibited by
(a) ATP
(b) Alanine (can be produced by pyruvate, so signals that other metabolic pathways are covered - similar to effect of citrate for above)
Activated by
(a) fructose 1, 6-bisphosphate (adapts rate of gylcolysis to the amount of substrate available)
Clinical features:
Inborn errors of carbohydrate metabolism
Hemolytic anaemia
Primary biliary cholangitis
Pyruvate kinase deficiency (results in anaemia with low hb levels - low hb levels less harmful than in other forms of anaemia as oxygen more readily released to cells)
B-Oxidation - Carnitine carrier
Role of fatty acids
Primary energy store, especially during prolonged fasting.
Mostly stored as triacylglycerols (TAG)/ triglycerides.
Preparation: release of FA from TAGS
Activation: forms a thioester with CoA in the cytosol - acyl-CoA.
Transportation: transported into the mitochondria via carnitine shuttle.
Degradation: in the mito, cleaved to Acetyl co-A (C2 SCFA) by Beta-oxidation.
Structure of Fatty acids
Fatty acids have a polar carboxylic acid “head” (COOH) and an apolar hydrocarbon “tail.” (Cn)
Saturated Fatty Acids: No double bonds. Example: Palmitic acid (C16).
Unsaturated Fatty Acids: Contain one or more double bonds. Double bonds indicated by delta.
Breakdown differs from saturated to unsaturated fatty acids.
Fatty acid oxidation
Oxidation - add oxygen - tends to be done by adding a phosphate group.
FA are activated by forming a thioester at the head group. This consumes ATP.
Coenzyme A (CoA) bonds via a suphide ion to the FA head.
Therefore called Acyl-CoA.
(Thioesters are high-energy compounds that contain sulfur and are more reactive than esters.)
Acyl-CoA vs Acetyl-CoA
Acyl-CoA: activated FA, includes FA of different chain lengths and saturation
Acetyl-CoA: specifically acetic acid activated with coenzyme A
Acetic acid is the main short-chain fatty acid (SCFA) (C2 unit) produced in the body, accounting for 50-70% of SCFA. It is the only SCFA that enters the bloodstream in significant amounts and can provide energy for muscles and other tissues - produced in glycolysis and fatty acid degradation, then further broken down in citric acid cycle.
What is Beta-oxidation?
Beta-oxidation refers to the oxidation of the beta carbon (C3) in the fatty acid chain.
Involves 4 steps.
(1) Oxidation: Acyl-CoA dehydrogenase converts the single bond between C2 and C3 into a trans double bond, forming delta-2-trans-enoyl-CoA. Electrons are transferred from the fatty acid to FAD, producing FADH2. (Energy balance +FADH2)
(2) Hydration: Enoyl-CoA hydratase adds a water molecule, converting the double bond back into a single bond and forming beta-Hydroxyacyl-CoA.
(3) Oxidation: 3-hydroxyacyl-CoA dehydrogenase oxidizes the hydroxyl group at the beta-carbon into a keto group (−C(=O)), producing beta-Ketoacyl-CoA and transferring electrons to NAD+, forming NADH and H+. (produces energy; Energy balance now +FADH2, + NADH plus H+)
(4) Thiolysis: Thiolase cleaves the fatty acid, releasing acetyl-CoA and shortening the acyl-CoA by two carbons. Keep entering Beta oxidation until only 2 C atoms left in 4th step.
Energy Yield: Each cycle yields one FADH2 (1.5 ATP) and one NADH plus H+ (2.5 ATP) . No direct ATP acquired by equates to 4 ATP per cleaved two-carbon unit in mitochondria (reduced by the electron chain)
***However, this only applies in the mitochondria - peroxisomal beta-oxidation produces less energy as FADH2 is regenerated without entering the electron transport chain. But still get NADH plus H+ (2.5 ATP).
Special Cases:
(1) Odd-Numbered Fatty Acids: Yield a three-carbon unit, propionyl-CoA, which converts into succinyl-CoA for the citric acid cycle.
(2) Unsaturated Fatty Acids: Require conversion of cis double bonds to trans double bonds to proceed with beta-oxidation.
Clinical Relevance: Disorders in beta-oxidation, such as MCAD, VLCAD, or LCHAD deficiencies, are rare and typically manifest during fasting due to glucose deficiency. They require dietary management, including carbohydrate-rich diets and avoiding fasting. Testing for these conditions is included in newborn screening.
What is the carnitine shuttle?
Activated fatty acids (acyl-CoA) must cross the mitochondrial inner membrane for degradation.
The carnitine shuttle facilitates this:
(1) CoA is replaced with carnitine in the cytosol (forming acylcarnitine).
(2) Acylcarnitine is transported into the mitochondrial matrix via carnitine-acylcarnitine translocase.
(3) Carnitine is replaced with CoA again inside the matrix to regenerate acyl-CoA.
See pic in folder
Primary carnitine def: extremely low levels of ketone bodies and glucose - develop hypoketotic hypoglycemia - failure to thrive, hypotonia, cardiomyopathy. PO supplementation.
