Glycolysis I lecture PDF

Preview

Summary

Dr. Potenza...

Description

Biochemistry Glycolysis I • Lecture 24 • • Lehninger Chapter 14 pp 533-553 • BCHE 395/451 October 17, 2018

1

Catabolism: The BIG Picture Carbohydrates

Proteins Fats

(1)

He ses

Amino acids Fatty acids

Pyruvate

α-keto acids Acetate

(2) (3) (4)

CO2

Krebs cycle

O2

NADH ADP

ATP

2

Central Importance of Glucose • 1) Glucose is an excellent fuel – Yields good amount of energy upon oxidation – Can be efficiently stored in the polymeric form – Many organisms and tissues can meet their energy needs on glucose only

• 2) Glucose is a versatile biochemical precursor – Bacteria can use glucose to build carbon skeletons of: • • • •

All the amino acids Membrane lipids Nucleotides in DNA and RNA Cofactors needed for the metabolism

3

Glucose oxidation via glycolysis

4

Four Major Pathways of Glucose Utilization • Storage – Stored as polymeric form (starch, glycogen) when there’s plenty of excess energy (é[ATP]&[NADH])

• Pentose Phosphate Pathway – Generates NADPH via oxidation of glucose for detoxification and the biosynthesis of lipids and nucleotides (ribose)

• Synthesis of Structural Polysaccharides – For example, in cell walls of bacteria, fungi, and plants

• Glycolysis – Generates energy via oxidation of glucose – Short-term energy needs 5

• Four total processes in breakdown of glucose • 1) Glycolysis: anaerobic breakdown of glucose (C6) to pyruvate (2 X C3) (cytosol) • 2) PDC/CAC: oxidation of pyruvate (C3) to CO2 + H2O + NADH/FADH2 • 3) Electron transport: conversion of NADH + ½O2 + H+ à H2O + NAD+ • 4) Oxidative phosphorylation: ADP + Pi à ATP

mitochondria

Glycolysis

6

Glycolysis • (1) Anaerobic metabolism: ààà à pyruvate glucose àà – D-Glucose + 2 ADP + 2 Pi + 2 NAD+ à 2 pyruvate + 2 ATP + 2 NADH + 4 H+ + 2 H2O

• Ubiquitous pathway for most organisms, both anaerobe and aerobes • Can occur in the absence of oxygen (ancient) • Energy yielding (+ 2ATP + 2NADH) • First metabolic pathway elucidated – 1930s: Germans Embden and Meyerhof à muscle 7

Glycolysis: Function • Pathway comprised of ten (10) total steps – No oxygen used in this initial oxidation

• Glucose is “activated” – (use 1 ATP) à Allows cell to extract MORE energy later

• Function: Glycolysis readies the molecule for further aerobic oxidation and energy extraction in later processes 8

Chemical Logic of Glycolysis

9

Chemical Logic of Glycolysis

“traps”

Highlighting transformations of carbon skeleton

Read and understand “The Importance of Phosphorylated Intermediates” pg 537-538 10

Glycolysis TP (- ) A

• Two-phases: • (1) Preparatory phase – Also, the “Investment” stage

• (2) Payoff phase – Also the “Dividend” stage burn 4 ATP Net = 2

(+) ATP (+) NADH

Investing 2 ATPs

“Lysis” Step Step 5 à All GAP Common product 12

How to Study Metabolic Pathways: •(1) Define pathway à Glycolysis (anaerobic process/oxidative): •Occurs in almost every living cell •Splits glucose (C6) into two C3 pyruvate units •Catabolic process: captures some energy (~5%) as 2 ATP and 2 NADH 13

How to Study Metabolic Pathways: •(2) Pathway Chemical summary equation: D-Glucose + 2 ADP + 2 Pi + 2 NAD+ à 2 pyruvate + 2 ATP + 2 NADH + 4 H+ + 2 H2O Energy made or used

14

How to Study Metabolic Pathways:

•(3) Look at each step for this information: •Rationale/logic •Substrates and products (equation): include e.g. ATP, NADH, CO2, H2O… •Type of reaction (1-6) and Mechanism •Enzyme: regulation, cofactors/coenzymes •Location of reaction step (cytosol? mitochondria?) •DG’° à favorable, unfavorable, reversible, irreversible, coupled? •Also check out DG for cellular conditions! 15

Step 1: Phosphorylation of Glucose 1st Priming Reaction This step uses 1 ATP/glucose TRAP

Addition of ℗

16

Step 1: Phosphorylation of Glucose glucose + ATP à Glu-6-P + ADP

• Rationale – Traps glucose inside the cell – Lowers intracellular glucose – Increases inherent free energy of product • Process uses energy of ATP • Hexokinase (eukaryotes): gluco (prokaryotes) • Mechanism: • ATP-bound Mg++ helps by shielding negative charges on ATP • Highly thermodynamically favorable/irreversible 17

Enzyme

Step 1: Hexokinase • Kinases associated with phosphoryl transfer and ATP; ‘hexo’ = 6 (carbons) – Enzyme that catalyzes transfer of phosphoryl group from ATP to a specified molecule

• Hexokinase (and other kinases) Mg2+ dependent • Regulated mainly by substrate inhibition • Widely distributed in nature; not specific to pathway, not specific to glucose • Isoenzymes: have distinctly different primary structure, electrophoretic, physical and chemical properties, but catalyze the same reaction 18

Step 2: Phosphohexose Isomerization

the arrows for the reaction: 2 way street because deltaG'knot is "neutral"

deltaG = -2.5 kJ/mol in cell 19

Step 2: Phosphohexose Isomerization • Rationale – Can’t phosphorylate aldehyde à 1C=O – C1 of fructose easier to phosphorylate by PFK (next enzyme) – Allows for symmetrical cleave by aldolase • Mechanism: aldose (glucose) isomerizes to a ketose (fructose) via enediol intermediate • Isomerization catalyzed by active-site glutamate à general acid/base catalysis • Slightly thermodynamically unfavorable/reversible – Product concentration kept low to drive forward – DG à cellular conditions 20

2nd Priming Phosphorylation

Step 3: Commitment 2nd priming reaction

The first committed step of glycolysis delta G'knot = kJ/mol

Step uses 1 ATP/glucose large negative deltaG'knot rxn direction

(FBP) product specifically targeted to glycolysis 21

Step 3: 2nd Priming Phosphorylation

fru 6-P + ATPà fru 1, 6 bis-P • Rationale

– Further glucose activation – 1 phosphate/3-carbon sugar after step 4 • First Committed Step of Glycolysis – fructose 1,6-bisphosphate committed to become pyruvate and yield energy

• Uses energy of ATP • Highly thermodynamically favorable/irreversible • highly regulated/allosteric – ATP, fructose-2,6-bisphosphate, other metabolites

– Why? Don’t ‘burn’ glucose if there is plenty of ATP (high energy charge) !!! 22

Enzyme

Step 3: Phosphofructokinase-1 • • • • •

Irreversible reaction (nearly!) First unique enzyme reaction in glycolysis Enzyme subject to STRONG metabolic regulation PFK-1 inhibited by ATP (and citrate…later…) Rate limiting reaction of glycolysis

23

Phosphofructokinase-1: Regulation • (1) Inhibited by high [ATP]

K0.5 change

– ATP is a reactant

• (2) Inhibition by ATP is reversed by AMP, ADP, Pi, or F-2,6-BP • Classic “R” enzyme • Allosteric

Catabolic Pathways Regenerate ATP

R

Rate of Reactions or Pathways U

[ATP] in cell

Anabolic Pathways Utilize ATP

24

25

Step 4: Aldol Cleavage of F-1,6-bP

C6 to C3

BUT ΔG in cell = -1.3 kJ/mole

Reverse aldol condensation (rxn of an alchohol and aldehyde) F-1,6-bP splits into 2 26

Aldolase ℗

Chpt 14 Q 6,9,20

1 2 3 4 5

℗

6

27

Step 4: Aldol Cleavage of F-1,6-bP • Rationale – Cleavage of a six-carbon sugar into two threecarbon sugars – High-energy phosphate sugars are three-carbon sugars (!) à triose phosphates • Reverse process is the familiar aldol condensation • Mechanism: covalent catalysis • Thermodynamically unfavorable/reversible – GAP concentration kept low to pull reaction forward – DG à cellular conditions 28

Step 5: Final Prep Step ketone to aldehyde Triose Phosphate Interconversion DHAP

Equilibrates DHAP and G3P Or makes them equivalent

(3, 4)

GAP G3P

(2)

(2, 5)

50:50 mix

(1)

(1, 6)

Notes: ΔG is negative in cell because [GAP] is low Also a enedio intermediate

Removal of GAP from the pathway via stage 2

(3)

29

Step 5: Triose Phosphate Interconversion • Rationale: – Allows glycolysis to proceed by one pathway • Aldolase creates two triose phosphates: – Dihydroxyacetone Phosphate (DHAP) – Glyceraldehyde-3-Phosphate (GAP)

• Mechanism is similar to phosphohexose isomerase: Just do it yourself • Only GAP is the substrate for the next enzyme (step 6) • DHAP must be converted to GAP • Completes preparatory phase • Thermodynamically unfavorable/reversible – GAP concentration kept low to pull reaction forward 30...