Outline Fate of pyruvate (summary) Energy yield of glycolysis

– Anaerobic– Aerobic

Details of the fates of pyruvate– Reduction of pyruvate to lactate– Oxidative decarboxylation of pyruvate to acetyl CoA– Carboxylation of pyruvate to oxaloacetate– Reduction of pyruvate to ethanol

Feeder pathways for glycolysis– Fructose– galactose

Fate of Pyruvate Lactate dehydrogenase

– RBC and during exercise– Reversible in liver– Location: cytoplasm

Pyruvate dehydrogenase– TPP, LA are cofactors– Source of AcetylCoA– Irreversible reaction– Location: Mitochondria

Ethanol synthesis– In yeast, some bacteria– Location cytoplasm

1. Reduction of pyruvate to ethanol (microorganism)

• It occurs by the 2 reactions shown below:

Pyruvate decarboxylase mechanism

• There is no net oxidation-reduction in the conversion of Glc into Ethanol.

• NAD+ is used first and made it later!

Active site of Alcohol dehydrogenase

Pyruvate decarboxylase is present in brewer’s and baker’s yeast. CO2 produced during alcohol fermentation is responsible for the characteristic carbonation of champagne.

In baking, CO2 fermentation by pyruvate decarboxylase during fermentation of dough due to CO2 , dough rises.

Alcohol dehydrogenase metabolizes alcohol.

TPP carries “active aldehyde” groups

The pyruvate decarboxylase reaction is the first reaction we see that TPP is involved. TPP Vit B1. If B1 is not enough

More about TPP

Beriberi: swelling, pain, paralysis, death

TPP plays an important role in the cleavage of bonds adjacent to a carbonyl group such as the decarboxylation of alpha-ketoacids and in chemical rearrangements involving transfer of an activated aldehyde group from one C to another.

The functional part of TPP is the thiazolium ring. The proton at C-2 of the ring is relatively acidic. Loss of this proton produces an active site in TPP.

TPP is involved in the following reactions1. Pyruvate decarboxylase2. Pyruvate dehydrogenase3. Alpha-ketoglutarate dehyrogenase4. Transketolase

Microbial fermentation yield other end products of commercial value:

Lactate and ethanol are the common products of microbial fermentation

Not the only ones...– In 1910, Chaim Weizmann (first president of Israel)

discovered a bacterium– Clostridium acetobutyricum ferments starch to

butanol and acetone. Here comes industrial fermentation, purpose is

to make important products from readily available material (like starch) by using microorganism.

Oxidation of ethanol in humans

Alcohol is a significant source of calories in many individuals. The metabolism of ethanol yields acetate by means of a pathway of 2 oxidation reactions.1. Formation of acetaldehyde:

– The first oxidation in the metabolism of ethanol occurs in the liver by alcohol dehydrogenase.

– Some acetaldehyde is formed by a microsomal ethanol oxidizing system (MEOS) involving NADPH, O2 and cyt P450.

2. Formation of acetate: – Acetaldehyde is further oxidized to acetate by the enzyme

aldehyde dehydrogenase.

Some information about alcoholism

90% people drink alcohol.

40-50% of male have temporary alcohol-induced problems.

10% male and 3-5% female have persistent alcohol related problems (alcoholism).

Ethanol easily moves through cell membranes.

12 oz beer or 4 oz wine has 10 grams of ethanol

1 liter of wine 80 g ethanol.

More about alcohol

It is a CNS depressant, decreases activity of neurons.

It is absorbed in the mouth, esophagus, stomach, and large bowel with a major site of absorption is small intestine.

The rate of absorption is increased with rapid gastric emptying, absence of proteins, fats, or carbohydrates.

2%-20% of ethanol is excreted directly through the lungs, urine and sweat.

The rest is metabolized to acetoaldehyde.

The clinical significance of acetaldehyde is not known, but accumulation in liver, brain, or other body tissues may cause organ damage.

2. REDUCTION OF PYRUVATE TO LACTATE:

Lactate formed by the action of lactate dehydrogenase is the final product of anaerobic glycolysis.

In exercising skeletal muscle, NADH production (by glyceraldehyde3-P and by the 3 NAD-linked dehydrogenase reaction of the TCA cycle) exceeds the capacity of the respiratory chain, resulting in an increase in NADH/NAD that favors reduction of pyruvate to lactate.

Therefore, during intense exercise, lactate accumulates in the muscle , pH decreases resulting in cramps. Much of the lactate eventually diffuses from the muscle into the blood stream.

The direction of lactate dehydrogenase reaction depends on the relative intracellular concentrations of pyruvate and lactate and on the ratio of NADH/NAD in the cell.

When animal tissues can not be supplied with enough oxygen to support aerobic oxidation of pyruvate and NADH produced in glycolysis, NAD+ is regenerated from NADH by the reduction of pyruvate to lactate.

More about lactate formation

Certain other tissues and cell types (such as retina, brain, and RBCs) produce lactate from pyruvate under aerobic conditions.

Lactate is the major end-product of glycolysis in RBCs.

In tissues such as the liver and heart, the ratio of NADH/NAD is lower than in exercising muscle. These tissues can oxidize lactate to pyruvate.

The cycle of reactions that includes glucose lactate in muscle and lactate glucose in liver is called the Cori Cycle for Carl and Gerty Cori whose studies in the 1930s and 1940s clarified the pathway and its role.

3. The oxidative decarboxylation of pyruvate:

The oxidative decarboxylation of pyruvate by pyruvate dehydrogenase is an important pathway.

Pyruvate + NAD+ CoA Acetyl CoA +CO2 + NADH

This goes to TCA cycle and will be discussed later.

ENERGY YIELD FOR GLYCOLYSIS

A. ANAEROBIC GLYCOLYSIS:

Overall reaction:

Glucose + 2P + 2ADP 2 Lactate + 2ATP + 2H2O

1. A net of 2 mols of ATP is produced for each molecule of glc.

2. Anaerobic glycolysis, although releasing only a small fraction of the energy contained in the glucose molecule, is a valuable source of energy under the following conditions.– When oxygen is decreased such as during intense exercise

– In tissues with few mitochondria – the kidney, RBCs, WBCs, retina, and brain.

3. In anaerobic glycolysis there is no net production or consumption of NADH. The NADH formed by glyceraldehyde 3-P dehydrogenase is used by lactate dehydrogenase to reduce pyruvate to lactate. Two trioses are produced for each glucose molecule.

ATP consumption and production

Reaction Change in ATP per Glc consumed

1. Glc-----> Glc-6-P …………..

2. Fructose-6-P------->F1,6bisP …………. 3. 1,3 bisPglycerate--------> 3-Phosphoglycerate ………….

4. PEP---------> Pyruvate ………….

Net: ………….

Formation and consumption of NADH in anaerobic glycolysis:

Reactionchange in NADH

1. Glyceraldehyde 3P-----> 1,3 BisPglyceraldehyde ……………

2. Pyruvate---------------> Lactate ……………

Net NADH: ……………

Aerobic glycolysis:

Overall reaction:

Glc+ 2P + 2NAD+ + 2ADP 2 Pyruvate + 2ATP + 2NADH + 2H+ + H2O.

1. The direct formation and consumption of ATP are the same as in anaerobic glycolysis, that is a net gain of 2ATP per molecule of glucose.

2. 2 mols of NADH are produced per mol glucose. Ongoing aerobic glycolysis requires the oxidation of this NADH by the respiratory chain.

Formation of ATP in aerobic glycolysis

Reaction Change in ATP/glc cons.

Glc-------------> Pyruvate

NADH--------> NAD+

Net ATP:

Other monosaccarides can enter the glycolytic pathway

Fructose must be phosphorylated to enter glycolysis.

2 enzymes are responsible for this step.

a. Hexokinase has a low affinity for fructose (high Km). Unless fructose levels of the body increases, little fructose is converted to fructose-6-P by this enzyme. D-fructose (present in fruits) or hydrolysis of sucrose can be phosphorylated by hexokinase to F-6-P.

Sucrose + H2O Fructose + Glucose

Fructose + ATP F-6-P + ADP

In muscle and kidney, this is a major pathway.

Second enzyme

b. By fructokinase:

The liver (which processes much of the dietary fructose) and the kidney have fructokinase which converts fructose to fructose-1-P. Fructose-1-P yields glyceraldehyde and DiOHacetoneP by aldolase B enzyme action. Two steps are bypassed (hexokinase and PFK) and the rate of fructose metabolism is greater than that of Glc.

Galactose metabolism: Dietary source is lactose from milk or milk products.

Lactose Glc + Galactose

Dietary disaccarides are hydrolyzed to monosaccarides by enzymes lining the small intestine.

– Maltose 2D-Glc

– Lactose D-Glc + D-Gal

– Sucrose > D-Glc + D-Fructose

– Trehalose 2D-Glc

Galactose metabolism

Gal Glc-6-P in four steps

1. Gal Glc interconversion pathway

2. Gal-1-P then takes uridyl group

3. UDP-Gal’s Gal is then epimerized to Glc

4. Finally, Glc-1-P isomerized to G-6-P

Many adults are intolerant of milk due to lactase deficiency

Lactose intolerance or hypolactasia– Deficiency is not quiet the appropriate term– Lactase activity declines to 5 to 10% of the level of birth– Most Africans and almost all Asians have very low levels!– Populations with a tradition of herding cattle (northern

Europeans) continue to express lactase gene Problem is diarrhea. Treatment is easy now: Milk products (lactose has

been hydrolyzed enzymatically) and lactase-containing pills are widely available.

Galactose is highly toxic

Galactosemia: Gal-1-P-uridyl transferase deficiency– Fail to thrive– Vomit– Jaundice– Cirrhosis– Cataracts

Blood Gal is high. The absence of the transferase in RBCs definitive

diagnosis. Why cataracts?

BSO+NACA

Grade 0

BSO only (grade 3)

Control (Grade 0)

NACA only (Grade 0)

REGULATION OF CARBOHYDRATE CATABOLISM

Carbohydrate catabolism provides: ATP precursors, building blocks of some other

biosynthetic processes

ATP level for a cell should be almost constant.ATP production ATP consumption

balance

We undergo lots of changes which affect metabolism such as:

increased muscular activity decreased oxygen availability decreased carbohydrate intake

more

Events alter ATP production and utilization. Since ATP levels should be kept almost constant, we need to regulate some enzymes in glycolysis pathway.

There are 4 enzymes that play a role in this regulation (in liver and muscle)

1. Glycogen phosphorylase, hormonal, allosteric, Ca++

2. Hexokinase3. PFK-14. PK

Hexokinase

Muscle hexokinase is inhibited by Glc-6-P. Whenever Glc-6-P is increased, this enzyme is inhibited.

Glucokinase is an isozyme of hexokinase (also called hexokinase-D) – Isozymes are different proteins that catalyze the

same reaction.

PFK-1 is under complex allosteric regulation

MOST IMPORTANT CONTROL POINT

F-6-P + ATP F1,6 bisP + ADP

• ATP• pH• Citrate (key intermediate of TCA cycle)• Fructose 2,6 bisphosphate

PK Increased ATP concentrations inhibit PK

allosterically by decreasing its affinity to its substrate. PEP

PK is also inhibited by Acetyl-CoA.

PEP Pyruvate Acetyl-CoA

Several different forms exist (L and M)• F 1,6 bisP activates PK to keep pace with oncoming

high flux of intermediates.• ATP inhibits, enough energy!• Alanine also inhibits PK• Reversible phosphorylation of the enzyme also

controls this enzyme’s action.

Family of Glc transporters

Common structure:– 12 transmembrane segments

The members of this family have some distinctive roles:

• glut1 and glut3• glut2• glut4• glut5

Cancer and glycolysis

Tumors have high rate of glc uptake and glycolysis

Why?

Hypoxic area in tumors

Consequences of the adaptation