Lec21/22 - An Overview of Further Metabolism (Carbohydrate Anabolism + Storage; Glucose Metabolic Regulation; Nitrogen Fixation; Amino Acid Metabolism; Lipid Metabolism) Flashcards
Define Gluconeogenesis
The synthesis of glucose from non-carbohydrate precursors, (e.g., alanine and lactate) via pyruvate
(Sort of inverse of glycolysis, but different enzymes for steps 1, 3 and 10)
What is the purpose of gluconeogenesis?
To maintain glucose levels in the blood and provide energy to the brain and muscles
Describe how step 1 of gluconeogenesis differs from step 10 of glycolysis
IN GLUCONEOGENESIS 1:
Pyruvate -> Oxaloacetate -> Phosphoenolpyruvate (catalysed by pyruvate carboxylASE, then phosphoenolpyruvate carboxylkinASE)
IN GLYCOLYSIS 10:
Phosphoenolpyruvate -> Pyruvate (Pyruvate KinASE)
Describe how step 8 of gluconeogenesis differs from step 3 of glycolysis
IN GLUCONEOGENESIS 8:
Fructose 1,6-bisphosphate -> Fructose 6-phosphate (catalysed by Fructose 1,6-bisphosphatASE)
IN GLYCOLYSIS 3:
Fructose 6-phosphate -> F1,6P (catalysed by phosphofructokinASE-1)
Describe how step 10 of gluconeogenesis differs from step 1 of glycolysis
IN GLUCONEOGENESIS 10:
Glucose 6-phosphate -> Glucose (Glucose 1,6-phosphatASE)
IN GLYCOLYSIS 1: Glucose -> Glucose 6-phosphate (HexokinASE, GlucokinASE)
Describe how the 4 (mentioned) non-carbohydrate precursors can enter the gluconeogenesis pathway
- Lactate <-> pyruvate
- Alanine <-> pyruvate
- Aspartate <-> oxaloacetate
- Glycerol/Fats <-> Dihydroxyacetone Phosphate
State the overall stoichiometric equation for Gluconeogenesis (and state what this shows about free energy compared to glycolysis)
2Pyruvate + 2NADH + 4ATP + 2GTP + 6H2O + 2H+
——>
Glucose + 2NAD+ + 4ADP + 2GDP + 6Pi
(More free energy to generate glucose from pyruvate [4+2 = 6ATP/GTP] than can be generated from glycolysis [2ATP])
What is pyruvate carboxylase?
An important enzyme at the start of gluconeogenesis, which catalyses a metabolically IRREVERSIBLE reaction that occurs in the mitochondrial matrix
State the equation of the reaction catalysed by Pyruvate Carboxylase
Pyruvate + CO2 + ATP -> Oxaloacetate + ADP + Pi
State the role of Biotin in Gluconeogenesis
It is a prosthetic group that is covalently attached to Pyruvate Carboxylase (via a Lys side chain) - the first enzyme in gluconeogenesis
It is a carrier of activated CO2 (carboxybiotin)
How is Pyruvate Carboxylase allosterically activated, and what is the advantage of this?
Allosterically activated by Acetyl CoA -> Accumulation of Acetyl CoA signals that energy is abundant, so gluconeogenesis is more useful than glycolysis
What is starch and where is it stored?
It is the glucose storage molecule in animals (equivalent to starch in plants) and is stored in cytosolic granules in the liver and muscle cells of vertebrates
What is Glycogenolysis, and what are the steps?
The degradation of glycogen:
Glycogen -> Glucose-1-phosphate -> Glucose-6-phosphate (PHOSPHOGLUCOMUTASE) -> Glucose (GLUCOSE-6-PHOSPHATASE)
How does glycogen compare to fatty acids as an energy store?
It is less reduced, so not as rich in energy - is is a quick source of glucose for a sudden demand in energy (also, released glucose can provide energy in the absence of oxygen)
How are glucose molecules removed from the glycogen polymer?
GLYCOGEN PHOSPHORLASE catalyses sequential removal of G1P (then the other enzymes convert this to glucose)
Describe what happens in glycogenolysis near a branch point in the molecule
Glycogen phosphorylase stops at 4 residues from a branch point - “limit dextrin” is then remodelled by transferase and a-1,6-glucosidase
Describe the role of transferase in glycogenolysis
At a branch point, transferase shifts three glucoses from one outer branch to the other
Describe the role of a-1,6-glucosidase in glycogenolysis
After transferase shifts 3 glucoses, a-1,6-glucosidase removes the single remaining branched glucose, leaving an elongated unbranched chain so that Glycogen Phosphorylase can continue
Summarise the process of glycogen synthesis
In the liver, cellular glucose is converted to G6P by GLUCOKINASE (using ATP)
Then one G6P molecule is incorporated into glycogen (using UTP)
Describe the process of UDP-Glucose (active form of glucose) formation in Glycogen Synthesis
The enzyme UDP-Glucose Pyrophosphorylase liberates the outer 2 Phosphates from UTP as PPi;
UMP + G1P -> UDP-Glucose (+PPi)
Describe the overall energy requirement of glycogen synthesis
Overall, 2ATP required to incorporate one glucose molecule into glycogen (1 from glycokinase reaction [G6P] and 1 to form UDP-Glucose [ATP energetically equivalent to UTP])
Describe the principle of Glucose Polymerisation in Glycogen Synthesis
Glycogen synthase catalyses addition of one glycose to glycogen via formation of a 1,4-glycosidic bond
Describe the role and process of Priming in Glycogen Synthesis
Glycogen Synthase needs at least 4 glucoses to add to (a primer)
GlycogenIN initiates glycogen synthesis by polymerising the first 10-20 glucoses
UDP-G + (glucosyl)nGlycogenin -> (glucosyl)n+1-Glycogenin + UDP
FACT: Transfer of glucose to growing chain via 1,4 glycosidic bonds (linear growth) is a MAJOR REGULATORY STEP IN GLYCOGEN SYNTHESIS
Cool beans
Describe how glucose enters mammalian cells
Down a concentration gradient, via GLUTs (e.g., GLUT1/2/3/4) - a family of passive hexose transporters that facilitate glucose transport
Describe the different roles of GLUT1/2/3/4
GLUT1 + 3 = nearly all mammalian cells, constantly transport glucose into cell under normal conditions at almost constant rate
GLUT2: Liver + Pancreatic ß Cells; transport glucose when [Glucose] in blood is high -> Pancreas detects -> Insulin production
GLUT4: Muscle and fat cells, numbers increase rapidly when insulin is present
How does insulin stimulate increased numbers of GLUT4?
It binds to an extracellular insulin receptor (A tyrosine kinase), which recruits vesicles containing GLUT4 to the cell membrane
How is regulation of Glycolysis and Gluconeogenesis linked?
Co-ordinated pathways: When one is active, the other is relatively inactive (rate of Glycolysis controlled by [Glucose]; rate of gluconeogenesis controlled by [Glucose Precursors])
Describe the role of Fructose 2,6-bisphosphate in regulating Glycolysis and Gluconeogenesis
It is a potent ACTIVATOR of PHOSPHOFRUCTOKINASE-1 (regulatory enzyme in Glycolysis);
and an INHIBITOR of FRUCTOSE 1,6-BISPHOSPHATASE (an enzyme specific to gluconeogenesis)
Describe the synthesis / counter-conversion of Fructose 2,6-bisphosphate (and how these two enzymes are linked)
Synthesised by Phosphorylation of F6P by LIVER PHOSPHOFRUCTOKINASE-2 (PFK2);
Converted back to F6P by FRUCTOSE BISPHOSPHATASE-2 (FBPase2)
Both enzymes are on the same protein chain, controlled by phosphorylation of a single Ser residue (+ = FBPase2; - = PFK2)
How is Glucagon involved in regulation of liver glycolysis?
When blood [Glucose] low (e.g., after night fast), glucagon rises and leads to phosphorylation of PFK-2/FBPase2, activating FBPase2, so levels of F2,6BP decrease (as converted to F6P), so gluconeogenesis dominates over glycolysis
When blood [Glucose] high (e.g., after meal), glucagon levels decrease and insulin produced, so PFK2 is activated, so F6P converted to F26BP, so PFK1 activated and glycolysis dominates
Which 3 hormones “also” regulate glycogen metabolism?
Insulin, Glucagon and Epinephrine/Adrenaline
What is the effect of insulin on regulation of glycogen metabolism?
Insulin (produced by pancretic ß cells) increases rate of glucose transport into muscle and adipose tissue, stimulating glycogen synthesis in the liver
What is the effect of Glucagon on regulation of glycogen metabolism?
Glucagon (produced by pancreatic alpha cells in response to low [Glucose]) stimulates glycogen degradation
What is the effect of insulin on regulation of glycogen metabolism?
Adrenline/Epinephrine (produced by adrenal glands in response to sudden energy requirement) stimulates the breakdown of glycogen to glucose, and the release of this glucose into blood
Distinguish between essential and non-essential amino acids
Essential amino acids cannot be synthesised and must be obtained from the diet
Non-essential amino acids can be synthesised
Where does the nitrogen in amino acids “come from”?
Nitrogen fixation: atmospheric N2 is converted into ammonia (NH3) by diazotrophic (nitrogen fixing) bacteria e.g., Rhizobium
Give the equation of the reaction that takes place to fix nitrogen in Rhizobium, and which enzyme catalyses it
N2 + 8H+ + 8e- + 16ATP -> 2NH3 + H2 + 16ADP + 16Pi (NITROGENASE)
Note: this reaction needs ATP and reducing power
How is ammonia incorporated into biomolecules (i.e. in what form)?
In the form of ammonium ions NH4+
Describe the structure of Nitrogenase
A tetramer of 2alpha and 2 beta units, also contains a unique iron-molybdenum cofactor
How is ammonium incorporated into amino acids in microorganisms (main chain)?
NH4+ incorporated into alpha-ketoglutarate (citric acid intermediate) to form L-GLUTAMATE
Catalysed by GLUTAMATE DEHYDROGENASE (reversible in microbes, same enzyme only catalyses reverse in humans)
Note: Needs reducing power NAD(P)H
How is ammonium incorporated into amino acids in microorganisms (side chain)?
NH4+ incorporated into glutamate to form GLUTAMINE (catalysed by GLUTAMINE SYNTHETASE)
Note: Needs ATP
How are other amino acids besides Glutamate and Glutamine synthesised in microbes?
Transaminase reactions: the amino group of glutamate is transferred to alpha-keto acids, generating alpha amino acids
Where are transaminases found in humans?
In the liver
Note: they are a measure of liver function
Name two examples (mentioned) of transaminase enzymes
Alanine aminotransferase and aspartate aminotransferase (form alanine and aspartate respectively)
Name the 9 essential amino acids
Valine, Leucine, Methionine, Isoleucine, Lysine, Threonine, Histidine, Tryptophan, Phenylalanine
Which amino acids are part of the Glycolysis -> Pyruvate biosynthetic family?
Alanine, VALINE, LEUCINE
Which amino acids are part of the Glycolysis -> 3-phosphoglycerate biosynthetic family?
Serine (Cysteine, Glycine)
Which amino acids are part of the Citric Acid Cycle -> Oxaloacetate biosynthetic family?
Aspartate (Asparagine, Methionine, Lysine, Threonine [Isoleucine])
Which amino acids are part of the Citric Acid Cycle -> alpha ketoglutarate biosynthetic family?
Glutamine, Proline, Arginine
Which amino acids are part of the Pentose Phosphate Pathway biosynthetic family?
Phenylalanine, (Tyrosine), Tryptophan, Histidine
What are the three main categories of animals in terms of ammonia excretion?
Ammonotelic = excrete ammonia directly in water (marine)
Ureotelic = urea (terrestrial vertebrates)
Uricotelic = Uric Acid (birds, reptiles, insects)
What are the two initial steps of Amino Acid Degradation?
Reversal of Synthesis Reactions:
- TRANSAMINASES transfer the amino group to alpha-ketoglutarate, forming glutarate (and alpha keto acid)
- GLUTAMATE DEHYDROGENASE oxidises glutamate and removes NH4+, (thus also regenerating alpha ketoglutarate)
What happens in amino acid degradation after ammonia is formed?
The Urea Cycle (involving several nitrogenated metabolites) to form urea
Why are triacylglycerols so rich in free energy (38kJ/g)?
Long hydrocarbon chains are the most reduced form of carbon
How are Glycogen and Triglycerides different in terms of their stored energy?
Glycogen can supply ATP for muscle contraction for less than 1 hour;
Sustained work is fueled by Catabolism of TGs
How are triglycerides stored (and broken back down into Glycerol and FAs)?
Stored as lipid droplets in the cytoplasm of specialised fat cells (adipocytes)
Hormone-stimulated lipases break TGs into glycerol and fatty acids
Describe the process of glycerol metabolism in the liver
Converted to Dihydroxyacetone Phosphate (DHAP), which is then isomerised to glyceraldehyde-3-phosphate (an intermediate in glycolysis/gluconeogenesis)
How are fatty acids transported in the blood and into cells?
In blood bound to albumin; enter cells via Fatty Acid Transport Protein (FATP) and bind to Fatty Acid Binding Protein (FABP)
(because they are water-insoluble)
How are fatty acids activated on the Outer Mitochondrial Membrane (so they can be oxidised)?
By Acyl coA Synthetase - forming Fatty Acyl CoA (thioester bond):
Fatty Acid + ATP + CoA <-> Fatty Acyl CoA + (AMP+PPi)
Note: they still cannot cross the membrane, need acyl carnitine translocase
How are Fatty Acyl CoA transported across the Inner Mitochondrial Membrane?
They cannot cross it, instead they are translocated by Acyl Carnitine Translocase:
Acyl CoA transfers the Acyl group to Carnitine; Acyl Carnitine then crosses the membrane (via the Translocase enzyme) then transfers the Acyl group back to a CoA
Which enzymes catalyse the transfer of the Acyl group between Carnitine and CoA in Fatty Acyl CoA OMM transport?
In cytosol: Carnitine Palmitoyl Transferase I (CPTI)
In matrix: Carnitine Palmitoyl Transferase II (CPTII)
Describe how ß-oxidation of Fatty Acyl CoA occurs
In the matrix, four enzymatic steps convert 2 carbons from Fatty Acyl CoA to Acetyl CoA (equivalent to 10 ATP by entering TCA)
What would be the net ATP yield for Palmitoyl CoA (a 16 carbon Fatty Acyl CoA)?
8 Acetyl CoA x 10 = 80ATP
7 FADH2 x 1.5 = 10.5 ATP
7 NADH x 2.5 = 17.5
-2 for Palmitate Activation
=106 ATP
Where does Fatty Acid synthesis occur in mammals?
In the cytoplasm of liver and adipocytes
What is required for fatty acid synthesis?
ATP and reducing power (NADPH)
When does fatty acid synthesis occur?
When glucose is plentifyl, excess acetyl coA is produced by glycolysis and is used for FA synthesis
Which enzyme(s) catalyse Fatty Acid Synthesis?
Fatty Acid Synthase (consists of several enzymes in a single polypeptide chain)
How does Fatty Acid synthesis fundamentally work?
Chain elongation, by sequential addition of two-carbon units derived from Acetyl CoA
Describe the first step of Fatty Acid Synthesis (all deets)
Acetyl CoA + ATP + HCO3- -> Malonyl CoA + ADP + Pi + H+
Acetyl CoA carboxylase catalyses this irreversible, committed step
