Glycolysis/EMP Pathway

Glycolysis/EMP

Glycolysis degrade a molecule of glucose in a series of enzyme-catalyzed reactions to give/yield 2 molecules of three-carbon(C3) compound pyruvate of lower free energy. Free energy released from glucose is conserved (use to synthesis) in the form of ATP and NADH.

·       Glycolysis reactions are take place in cytosol.

Major contributors:

1.     Gustav Embden

2.     Otto Meyerhof

3.     Jacob Parnas

Chemical Strategy of Glycolysis is:

1. Add phosphoryl groups to the glucose
2. Chemically convert phosphorylated intermediates into compounds with high phosphate group-transfer potentials.
3. Chemically couple the subsequent hydrolysis of reactive substances to ATP synthesis.

Stages of Glycolysis

Glycolysis has 10 steps which can dived into 2 phases.

• ·       Stage I (Reactions 1–5) - A preparatory stage - hexose glucose is phosphorylated and cleaved to yield 2 molecules of the triose glyceraldehyde-3-phosphate. This process utilizes 2 ATPs in a kind of energy investment 

• ·       Stage II (Reactions 6–10): Payoff phase -

The 2 molecules of glyceraldehyde-3-phosphate are oxidative converted to pyruvate, with generation of 4 ATPs & 2NADH.so, net ATP gain per glucose molecule in glycolysis is 2.

extra -

Energy Remaining in Pyruvate

Glycolysis releases only a small fraction of the total available energy of the glucose molecule; the two molecules of pyruvate formed by glycolysis still contain most of the chemical potential energy of glucose.

    

Importance of Phosphorylated Intermediates

Each of the 9 glycolytic intermediates between glucose and pyruvate is phosphorylated. The phosphoryl groups appear to have 3 functions -

1. Because the plasma membrane generally lacks transporters for phosphorylated sugars, the phosphorylated glycolytic intermediates cannot leave the cell. After the initial phosphorylation, no further energy is necessary to retain phosphorylated intermediates in the cell, although the large difference in their intracellular and extracellular concentrations 
2. Phosphoryl groups are essential components in the enzymatic conservation of metabolic energy. (extra - Energy released in the breakage of phosphoanhydride bonds (such as those in ATP) is partially conserved in the formation of phosphate esters such as glucose 6-phosphate. High-energy phosphate compounds formed in glycolysis (1,3-bisphosphoglycerate and phosphoenolpyruvate) donate phosphoryl groups to ADP to form ATP)
3. Binding energy resulting from the binding of phosphate groups to the active sites of enzymes lowers the activation energy and increases the specificity of the enzymatic reactions. (extra - The phosphate groups of ADP, ATP, and the glycolytic intermediates form complexes with Mg²⁺ and the substrate binding sites of many glycolytic enzymes are specific for these Mg²⁺ complexes. Most glycolytic enzymes require Mg²⁺ for activity)

Reaction 1 -Phosphorylation of Glucose

• 1^st Reaction of glycolysis is the transfer of a phosphoryl group from ATP to glucose to form glucose-6-phosphate (G6P) in a reaction catalyzed by hexokinase (HK)
• kinases are the enzymes that transfers phosphoryl groups between ATP and a metabolite
• Hexokinase, like many other kinases, requires Mg²⁺ for its activity, because the true substrate of the enzyme is not ATP^4- but the MgATP^2- complex. Mg²⁺ shields the negative charges of the phosphoryl groups in ATP, making the terminal phosphorus atom an easier target for nucleophilic attack by an -OH of glucose

Reaction 2 - Conversion of Glucose 6-Phosphate to Fructose 6-Phosphate

• conversion of G6P to fructose-6-phosphate (F6P) by phosphoglucose isomerase (PGI; also called glucose-6-phosphate isomerase/ phosphohexose isomerase)
• This is the isomerization of an aldose to a ketose

Reaction 3 - Phosphorylation of Fructose 6-Phosphate to Fructose 1,6- Bisphosphate

• phosphofructokinase (PFK) catalyzes the transfer of a phosphoryl group from ATP to fructose 6-phosphate to yield fructose 1,6-bisphosphate(FBP) [previously known as fructose-1,6-diphosphate (FDP)]
• PFK plays a central role in the control of glycolysis because it catalyzes one of the pathway’s rate-determining reactions

Reaction 4 - Cleavage of Fructose 1,6-Bisphosphate

• Aldolase catalyzes the cleavage of FBP to form the 2 trioses glyceraldehyde-3-phosphate (GAP)and dihydroxyacetone phosphate (DHAP)
• Aldol cleavage between C3 and C4 of FBP requires a carbonyl at C2 and a hydroxyl at C4.

Reaction 5- Interconversion of the Triose Phosphates

• DHAP and GAP are ketose–aldose isomers
• Triose phosphate isomerase (TIM or TPI) catalyzes this process

Reaction 6- Oxidation of Glyceraldehyde 3-Phosphate to 1,3-Bisphosphoglycerate

• This involves the oxidation and phosphorylation of GAP to 1,3-bisphosphoglycerate by NAD⁺ and P_i as catalyzed by glyceraldehyde-3-phosphate dehydrogenase (GAPDH)
• aldehyde oxidation, an exergonic reaction, drives the synthesis of the acyl phosphate group at C-1 1,3-bisphosphoglycerate.

Reaction 7- Phosphoryl Transfer from 1,3-Bisphosphoglycerate to ADP

• phosphoglycerate kinase(PGK) transfers the high-energy phosphoryl group from the carboxyl group of 1,3-bisphosphoglycerate to ADP, forming ATP and 3- phosphoglycerate.

Reaction 8- Conversion of 3-Phosphoglycerate to 2-Phosphoglycerate

• phosphoglycerate mutase(PGM) catalyzes the transfer of phosphoryl group between C-2 and C-3 of glycerate which results in conversion of 3PG to 2-phosphoglycerate (2PG).(mutases catalyze the transfer of a functional group from one position to another on a molecule)

Reaction 9 - Dehydration of 2-Phosphoglycerate to Phosphoenolpyruvate

• 2PG is dehydrated to phosphoenolpyruvate (PEP)in a reaction catalyzed by enolase
• The enzyme forms a complex with a divalent cation such as Mg²⁺ before the substrate is bound

Reaction 10 - Transfer of the Phosphoryl Group from PEP to ADP

• Transfer of the phosphoryl group from phosphoenolpyruvate to ADP, catalyzed by pyruvate kinase, which requires K⁺ and either Mg²⁺ or Mn²⁺
• This is a substrate-level phosphorylation

Overall reaction of glycolysis -

Glucose +2NAD⁺ +2ADP +2P_i →

2 pyruvate +2NADH +4H⁺ +2ATP +2H₂O

Regulation of Glycolysis

Enzyme	Inhibitors	Activators
Hexokinase	G-6-P
Phosphofructokinase	ATP, PEP, Citrate	ADP, AMP, Fructose 2,6-P
Pyruvate kinase	ATP

The Oxidizing Power of NAD⁺ Must Be Recycled

• NAD⁺ is the primary oxidizing agent of glycolysis
• The NADH produced by this process must be continually re-oxidized to keep the pathway supplied with NAD⁺

There are three common ways that this occurs -

1. Under anaerobic conditions in muscle, NAD⁺ is regenerated when NADH reduces pyruvate to lactate.
2. Under anaerobic conditions in yeast, pyruvate is decarboxylated to yield CO₂ and acetaldehyde and the latter is reduced by NADH to yield NAD⁺ and ethanol.
3. Under ATPs aerobic conditions, the mitochondrial oxidation of each NADH to NAD⁺ yields 2.5

Thus, in aerobic glycolysis, NADH may be thought of as a “high-energy” compound, whereas in anaerobic glycolysis its free energy of oxidation is dissipated as heat.

References - 

1. Lehninger Biochemistry
2. BIOCHEMISTRY by VOET . D & VOET J.G
3. Bacterial Metabolism by Gerhard Gottschal 
4. MICROBIAL PHYSIOLOGY by Albert G. Moat , John W. Foster & Michael P. Spector

Special thanks - 

Dr. Gagandeep Kaur