Introduction to Metabolism

Carbohydrate Metabolism

• Readings in Lehninger, 5th edition, are relevant to this topic.

Intermediary Metabolism

• Intermediary metabolism encompasses all reactions involved in breaking down compounds.
• It also includes generating and storing energy for the cell and organism's needs.
• Reactions involved in producing compounds (metabolites) for the cell or organism are part of intermediary metabolism.

Introduction to Metabolism

• Complex substances are broken down to produce energy, required metabolites, and structural components.
• Cells synthesize new complex substances.
• Thousands of reactions occur simultaneously in a single cell.
• Reactions occur with minimal side products, energy loss, and at reasonable conditions.
• Temperature, pH, and pressure are kept at a physiological level.
• All reactions are controlled and regulated for optimum efficiency.

Definitions:

• Catabolism refers to the breakdown of complex substances.
• Anabolism refers to the synthesis of complex substances from simpler ones.

Free Energy Changes in Metabolism

• Overall change in free energy (ΔG) is negative for catabolic processes.
• Example: higher energy compound G1 becomes lower energy compound G2, making ΔG negative.
• For an anabolic process, ΔG is positive.
• Energy, often from ATP, must be supplied to drive anabolic processes and make the overall ΔG negative.
• Catabolic processes generally generate energy for anabolic processes.

General Pathways of Metabolism: Catabolism

• Macromolecules break down into building blocks via hydrolysis.
• Proteins break down into amino acids
• Polysaccharides break down into glucose and other sugars
• Lipids break down into glycerol and fatty acids
• Nucleic acids break down into ribose, nitrogenous bases, and phosphate
• This step doesn't yield usable energy, only building blocks.
• Monomers break down into common intermediates such as pyruvate and acetyl CoA.
• Amino acids, glucose, glycerol, and fatty acids are converted to pyruvate or acetyl CoA.
• Pyruvate and acetyl CoA go through the citric acid cycle, ETS/Ox Phos to produce ATP.
• Oxidative processes produce ATP and NADH for energy.
• Intermediates break down to CO2 and electrons through a central oxidative pathway.
• The Citric Acid Cycle (TCA or Krebs Cycle) is the central cycle.
• The cycle leads to ATP production through electron transport and oxidative phosphorylation.

Anabolism

• Critical common intermediates, including TCA cycle components are utilized to make building blocks.
• Synthesizing building blocks requires energy in the form of ATP.
• Synthesizing macromolecules requires energy in the form of ATP.
• CO2 is not generally reused

Nutrient Requirements

• Some cells have specific nutrient requirements and cannot make some compounds.
• Vitamins may need to be externally obtained
• Some amino acids (about half) are required in the diet by humans.
• Some microorganisms cannot make certain amino acids and vitamins.
• These amino acids/vitamins must be supplied in nature or in a growth medium.

General Principles of Metabolism

• The processes of metabolism are highly controlled.
• Anabolism and catabolism aren't necessarily balanced.
• One may predominate in certain cells or at different times depending on the cell's needs.
• The pathway to synthesize a complex substance isn't simply the reverse of the degradative pathway.

Modes of Control

• Level of energy: if energy is low, anabolism is unlikely or impossible.
• Level of substrates must be at appropriate level for reactions to proceed
• Level of enzyme cofactors: lipoic acid, thiamine, NAD+, etc. must be present
• pH affects ionization states, meaning a molecule may be reactive only if in a protonated or unprotonated state
• Enzymes.
  • Quantity: regulated by repression or induction of gene expression.
  • Activity: enzymes may have inactive or less active states.
    • Allosteric enzymes have + or - effectors.
    • Product build-up leads to feedback control, inhibiting the enzyme.
• Compartmentalization: enzymes and substrates are restricted to certain organelles.
  • Making the substrate and enzyme available together in the right place.
• Hormone Control:
  • Certain cells are targeted by hormones, which indirectly regulate cellular pathways.
  • Hormones are small regulatory molecules synthesized elsewhere and delivered to target cells.
  • One type of hormone regulates metabolism by affecting gene expression (e.g., steroids).
  • Another type regulates metabolism through a second messenger system.
  • Hormones act at the outside surface of the cell and cause changes in internal levels of small molecules.
  • Cyclic AMP (cAMP) indirectly modifies enzyme activities.

Carbohydrate Metabolism Overview

• Carbohydrate metabolism revolves around glucose.
• Glucose can be converted to or from glycogen, pentose, and other sugars.
• Glucose can undergo glycolysis to produce pyruvate, which can then be converted to lactate, acetyl CoA, or EtOH

Enzyme Classification

• Dehydrogenase: oxidizes a substrate using cofactors as an electron acceptor or donor (e.g., pyruvate dehydrogenase).
• Reductase adds electrons from some reduced cofactor (e.g., enoyl ACP reductase).
• Kinase phosphorylates a substrate (e.g., hexokinase).
• Hydrolase uses water to cleave a molecule.
• Phosphatase hydrolyzes phosphate esters (e.g., glucose-6-phosphatase).
• Esterase (lipase) hydrolyzes esters (those that act on lipid esters are lipases) example lipoprotein lipase.
• Thioesterase hydrolyzes thioesters.
• Thiolase uses thiol to assist in forming a thioester (β-ketothiolase).
• Isomerase interconverts isomers (example aldose to ketose) triose phosphate isomerase.

Glycolysis

• Glucose is converted to glucose 6-phosphate by hexokinase using ATP.
• Glucose 6-phosphate is converted to fructose 6-phosphate by phosphoglucose isomerase.
• Fructose 6-phosphate is converted to fructose 1,6-bisphosphate by phosphofructokinase using ATP.
• Fructose 1,6-bisphosphate is converted to dihydroxyacetone phosphate and glyceraldehyde 3-phosphate by aldolase.
• Dihydroxyacetone phosphate can be converted to glyceraldehyde 3-phosphate by triose phosphate isomerase.
• Glyceraldehyde 3-phosphate is converted to 1,3-bisphosphoglycerate by glyceraldehyde 3-phosphate dehydrogenase, producing NADH.
• 1,3-bisphosphoglycerate is converted to 3-phosphoglycerate by phosphoglycerate kinase, producing ATP.
• 3-phosphoglycerate is converted to 2-phosphoglycerate by phosphoglyceromutase.
• 2-phosphoglycerate is converted to phosphoenolpyruvate by enolase, releasing H2O.
• Phosphoenolpyruvate is converted to pyruvate by pyruvate kinase, producing ATP
• Pyruvate can undergo:
  • Alcohol Fermentation
  • Anaerobic Glycolysis
  • Aerobic Glycolysis

Glycolysis: What is it?

• Glycolysis is a ten-step metabolic pathway.
• Converts glucose into two molecules of pyruvate.
• Also produces two molecules each of NADH and ATP.
• All carbohydrates to be catabolized must enter the glycolytic pathway.
• Glycolysis is central to generating both energy and metabolic intermediates.
• Pyruvate can be anaerobically converted to lactate in muscle and certain micro-organisms.
• Pyruvate can be anaerobically converted to ethanol (fermentation).
• Pyruvate can be aerobically converted to CO2 and H2O via the citric acid cycle.

Efficiency of Glycolysis

• Alcohol fermentation and anaerobic glycolysis (lactate) are inefficient energy providers.
• The glycolytic pathway yields little ATP.
• Aerobic processing of pyruvate through the citric acid cycle and respiration gives more complete oxidation.
• Also yields much higher ATP.
• Glycolysis provides pyruvate
• Pyruvate is a precursor for respiration with oxygen.
• Glycolysis in two stages:
  • An energy investment phase (reactions 1-5).
    • Converts glucose to two glyceraldehyde 3-phosphate molecules
    • Two ATPs are invested.
  • An energy payoff phase (reactions 6-10).
    • Converts two glyceraldehyde 3-phosphate molecules to two pyruvate plus four ATP molecules.
    • Net of two ATP molecules overall plus two NADH.

Phase I: Energy Investment

• Glucose is phosphorylated.
• Glucose enters a cell through a specific glucose transport process.
• The transport process is quickly phosphorylates at the expense of an ATP.
• Investment of an ATP is called "priming.”
  • Enzymes involved include hexokinase or glucokinase

Reaction Specifics

• Reaction: first energy investment.
• Highly exergonic, AG°´ = -16.7 kJ/mole and is essentially irreversible.
• Hexokinase found in all cells of every organism.
• Has low specificity for monosaccharides (simple sugars).
• Other monosaccharides can be phosphorylated by hexokinase.
• Relatively high affinity for glucose, KM = 0.1 mM.
• Inhibited by its product, glucose 6-phosphate.

Glucokinase

• Found in liver.
• High KM (~10mM) for glucose.
• Not inhibited by glucose-6-phosphate.
• Most effective when the glucose level in blood is high, such as after a meal.
• Isomerization of glucose 6-phosphate
• Enzyme involved is phosphoglucoisomerase
• Aldose to ketose isomerization
• Reversible, ΔG°´= 1.7 kJ/mole

Phosphorylation and Enzymes

• Second phosphorylation is facilitated by the enzyme phosphofructokinase.
• The second ATP investment in glycolysis is a highly exergonic and essentially irreversible reaction, with ΔG°´ = -14.2 kJ/mole.
• The phosphorylation step is highly regulated to modulate carbon flux through glycolysis, responding to energy and carbon requirements
• Fourth: Cleavage to two triose phosphates
  • The cleavage is facilitated by aldolase
    • Aldolase cleaves a 6 carbon sugar into two 3 carbon sugars.
  • High and positive ΔG ( '+23.8 kJ/mole)
• Fifth: Isomerization of dihydroxyacetone phosphate
  • Reaction facilitated by triose-phosphate isomerase
    • Triose-phosphate is important for allowing interconversion of the two triose phosphate products of aldolase cleavage to take place.
    • Interconversion occurs because only glyceraldehyde with its phosphate can be used for glycolysis.
    • Allows dihydroxyacetone phosphate to be metabolized by reacting it similiarly to glucuonse and fructose to make isomerase.
  • ΔG = +7.5 in reversible conditions.

Second Phase

• End of First Phase:
• Production of two glyceraldehyde 3-phosphate molecules from one glucose molecule with the expenditure of two ATPs.
  • The energy that steps in phase two yields are multipled by two.
• 6- Oxidation of glyceraldehyde 3-phosphate, facilitated by glyceraldehyde-3-phosphate dehydrogenase
• addition of phosphate, oxidation, production of NADH, formation of high energy compound
• product is an acylphosphate, a fused carboxylic-phosphoric acid anhydrate, which has a very high free energy of hydrolysis.
• reversible rxn, AG°´ = +6.3 kJ/mole because this fused group retains some of the energy produced by the oxidation of the aldehyde to the carboxylic acid.
• reaction produces important reducing compound NADH nicotinamide adenine dinucleotide, reduced form.
• First high energy compound generated = beginning of payoff.
• Enzyme is recycled and not used up in metabolism
• 7- Transfer of phosphate to make ATP
• Enzyme = phosphoglycerate kinase
• First substrate level phosphorylation, yielding ATP
• 2 1,3 bis PG yield 2 ATPs, thus so far ATP yield = ATP input
• high free energy yield, AG°´ = -18.8kJ/mole drives several of the previous steps.
• 8- Phosphate shift setup
• Enzyme= phosphoglycerate mutase
  • shifts phosphate from position 3 to 2
  • reversible, AG°' = + 4.6 kJ/mole
• mechanism involves phosphorylated enzyme intermediate with the formation of 2,3 bisphosphoglycerate
• 9- Second Generation of very high energy compound by a dehydration
• Enzyme = enolase
• Enols have an energy change of: +1.7 kJ/mole because the energy is locked into enolphosphate
• the energy is locked into the high energy unfavorable enol configuration by phosphoric acid ester
• the phosphoate is released and this energy is recovered to aid glycolysis.

Important Notes for Glycolysis Accounting

• A total of 2 ATPs are invested to drive glycolysis
• 2 ATPs from each glyceraldehyde 3-phosphate = total of 4 per original glucose in second phase
  • Net +2 ATP
• 2 molecules of NADH are also produced.

Summary of Glycolysis

• Input = 2 ATP
  • glucose + ATP → glucose-6-P
  • fructose-6-P + ATP → fructose 1,6 bisphosphate
• Output = 4 ATP + 2 NADH
• 2 glyceraldehyde 3-P + 2 P₁ + 2 NAD+→ 2 (1,3 bisphosphoglycerate) + 2 NADH
• 2 (1,3 bisphosphoglycerate) + 2 ADP→ 2 (3-P-glycerate) + 2 ATP
• 2 PEP + 2 ADP → 2 pyruvate + 2 ATP
  • = 2 ATP and 2 NADH
• Energy Yield From Glycolysis produces more energy but to an insigifnant degree compared to the intial glucose:
• glucose→ \6 CO2 = -2840 kJ/mole
• 2 ATPs produced = 2 x 30.5 61 kJ/mole glucose
• Engery is too low to use

Fate of Glycolysis:

• It has products that are centrally in metabolic pathways,

Lactatae Fermentation And Enzyme Importance

• Pyruvate uses oxygen in aerobic conditions with the aide of dehydrogenase lactate enzymes:
  • to lactate via lactate dehydrogenase
  • to ethanol via ethanol dehydrogenase
    • Both processes NADH in order to consume the products.
• Lactate Fermentation
• Enzyme = Lactate Dehydrogenase helps the body make and replenish the products with the aide of oxygen.
  • Note: uses up all the NADH (reducing equivalents) produced in glycolysis as the reuction is complete.
  • This is important for regeneration an alternative metabolic step.
  • Lactate, parts of the retina, and in skeletal musele cells during exercise.
• This fermentation is more important in plants and in some mirobes without oxygen. _ Lactate Dehydrogenase (LDH) has multiple forms (isoenzyme). Two polypeptides M and H come together to form LDH. The mixture will be like a tetramer:
  • M4, M3H, M2H2, MH3 and H4

Location of Lactate

• Skeletal muscle and liver contain mainly M forms; cardiac tissue, H forms.
• During and after cardiac attack, cardiac cells die in circulation to indicate some problems and or stress. These stresses are good to evaluate during diagnostic actions.

The Liver and Muscle Tissue Exchange

• The liver uses most of this lactate to make glycogen. Only small amounts of free glucose released.
  • Only small amount of glucose needs to be relased
• Glycogen can be broken down into glucose when needed.
• However, its rare because glucose is used almost immediateley

Alchoholic Fermentation

• Has a similar action with lactate But in this mechanism the ethanoyl group of oxygen in dehydrogenate isirreversible In additon oxygen + NADH is used mainly and quickly Ethanol does what alcohaols use in humans

Summary

• Has: Glucose converted to 2 ATP 2 NADH Products result in 2 NADH anaerobic 2 ethanol + CO₂ Resulting in: acetyl CoA + 2 CO2 Also producing oxygen: This produces oxygen and allows areobic resperation with: 4 CO2 + 4 H2O

Overview On Resperation

• The regulation results in a balance between the the products and regulation.
• Glycolysis with high enymes activity and concentration with high hexo and glucokinases.
• Phosphorylation of glucose. Inhibited by its product, glucose 6-phosphate, as a response to Glycolysis.

The Regulators

• Glucohinase enzymes are always a part of the liver and as a tool with high potency of glucose.
  • Are enzymes sensitive that increase potency to the body
  • Are in the blood system and not inhibited by glucose 6-phosphate.

Potency factors of Regulation

Are always the body in potency if they are high

Regulatiion must have what it offers to the body to benefit it The messure of all this benefits must be accurate Inhibited energy must be always low or normaly Activators. Enzymes that act in potency must be in good conditions

Inhibiting Factors

They have potentic to prevent frcotis

• AMP
• ADP
• 6 phosphate from enzymes Are importnant regualtors that act on controlling the blood.

Important Factors

• Act on liver and blood to balance
• Balance body Are inactive for easy digestion and stress free body Also the body with high levle of gene expression is good for loading.