iii. metabolism glucose catabolism – part...

Biochemistry 3300 Slide 1

III. Metabolism

Glucose Catabolism – Part II

Department of Chemistry and BiochemistryUniversity of Lethbridge

Biochemistry 3300


Metabolic Fates of NADH and Pyruvate

Cartoon:

Fate of pyruvate, theproduct of glycolysis.

+O2 TCA cycle

-O2

Fermentation



Pyruvate is a central branch point inMetabolism.

Aerobic pathway:Citric acid cycle and then respiration;- yields far more energy (discussed later) than glycolysis

- relatively slow (limited by O2 transport)

NADH + O2 → NAD+ + Energy

Pyruvate + O2 → 3 CO2 + Energy



Pyruvate is a central branch point inMetabolism.

Two anaerobic pathways:-Pyruvate is converted to lactate via lactate dehydrogenase (ie. muscle cells)

-Pyruvate is converted to ethanol via ethanol dehydrogenase (ie. yeast)

Both pathways use the NADH (produced in glycolysis): Overall: Glucose → 2 lactate + 2 ATP

Anaerobic pyruvate utilization = Fermentation


Lactate Fermentation

Pyruvate + NADH + + H+ L-Lactate + NAD+

Enzyme = Lactate Dehydrogenase

Regenerates NAD+ from NADH (reducing equivalents) produced in glycolysis.Essential as NAD+ is required for glycolysis (step 6 -GAPDH)

Lactate fermentation is important in red blood cells, parts of the retina and in skeletal muscle cells during extreme high activity.

Also important in plants and microbes growing in absence of O2.

∆G’° = -25.1 kJ/mol


Lactate Dehydrogenase (LDH)

In mammals two different types of LDH subunits are found: the M type and the H type.

Five forms of the tetrameric isozymes are possible:

M4, M3H1, M2H2, M1H3, H4

H-type predominates aerobic tissues (ie. heart muscle) H4 LDH has a low KM for pyruvate and is allosterically inhibited by it.

M-type predominates in tissue subject to anaerobic conditions (ie. liver and skeletal muscle) M4 LDH has a low KM for pyruvate and is NOT allosterically inhibited by it.


Lactate Dehydrogenase (LDH)

NADH

LDH monomer

NADH shown as sticksCatalytic site circled

Redox reaction involvingelectron transfer fromNADH to pyruvate.


Reaction Mechanism of Lactate Dehydrogenase


Pyruvate: Terminal Electron Acceptorof Lactic Acid Fermentation

Corey Cycle:

Most lactate is exported from the muscle cell via the blood to the liver↓

Liver converts lactate (back) to glucose↓

Glucose is transported from liver cells via the blood to the muscle(stored as glycogen)

The process of transporting lactate to the liver and its conversion to glucose takes from hours to days to complete.

Fate of Lactate (from fermentation)


Alcoholic Fermentation

Two enzymes involved: Pyruvate decarboxylase irreversible

Alcohol dehydrogenase reversible

Pathway is active in yeast

Regenerates NAD+ from NADH (reducing equivalents) produced in glycolysis.

Second step is reversible Ethanol can be further metabolised via oxidation that ultimately produces acetate and enters fat biosynthesis pathways


Pyruvate Decarboxylase(Alcohol Fermentation)

Yeast produces CO2 and ethanol in two consecutive reactions

Decarboxylation of pyruvate to acetaldehyde is catalyzed bypyruvate decarboxylase (PDC) (not present in animals).

PDC contains a tightly non covalently bound coenzyme:

Thiamin pyrophosphate (TPP)

Catalytically active


TPP Cofactor (Pyruvate Decarboxylase)

Decarboxylation of α-keto acidsbuilds up negative charge on the carbonyl carbon.

Transition state is stabilized bydelocalization of the developingneg. charge into a “electron sink”.

The dipolar carbanion (ylid) is theactive form



Thiamine Pryophosphate(TPP)

TPP of Pyruvate Decarboxylase:

Two views related by 90º rotationabout a vertical axis



How is TPP deprotonated toits the ylid form?

1) TTP’s aminopyradine ring (subunit 1) is deprotonated by Glu51 (subunit 2) of the PDC dimer. 2) amine of aminopyradine deprotonates thiazolium ring producing ylid form TPP

Note: PDC is a dimer of dimers.Note: TPP ylid form circled in red



Glu51

Thiamine Pyrophosphate(TPP)


Thiamine Deficiency

TPP addition to carbonyl groups and its ability to act as an“electron sink”(electron withdrawl) makes it the coenzyme most utilized in α-keto acid decarboxylations.

Thiamin (vitamin B1) is not synthesized or stored in significant amounts by vertebrates. Deficiency in humans results in an ultimately fatal condition known as beriberi.


Alcoholic Fermentation (step II)

Reduction of acetaldehyde to ethanol and regeneration of NAD+

by alcohol dehydrogenase (ADH)

Each subunit of the tetrameric yeast ADH binds one NADH and one Zn2+.


Alcoholic Fermentation Part II

Zn2+ polarises the carbonyl oxygenof acetaldehyde

Hydride ion is transferred fromNADH to the carbonyl carbon

Reduced intermediate acquires aproton from the medium to formethanol.


Glycolysis: Substrates other than glucose

Glycogen / Starch

Dietary Polysaccharides Maltose (Glu-Glu) Lactose (Glu-Gal) Sucrose (Glu-Fru)


Feeder Pathways for Glycolysis

Glycogen metabolism

Glycogen storage granulesin liver

Enzymes of 'feeder pathways'are underlined in red


Phosphorolysis:glycogen / starch degradation

Glycogen phosphorylase / Starch phosphorylase

- attack of Pi on the (α1→4) glycosidic linkage of the last two glucose residues.

Phosphorolysis generatesG1P which must be converted to G6P (phosphoglucomutase)to enter glycolysis


Phosphorolysis: glycogen / starch degradation

Phosphorylase - repetitively breaks (α1→4) linkages until it reaches an (α1→6)- produces glucose-1- phosphate

Debranching enzyme - required to break (α1→6) linkages - produces glucose


Phosphoglucomutase mechanism

Glucose 1-phosphatehas to be converted into glucose 6-phosphateto enter glcolysis

Where have we previouslyseen this type of mechanism?


Phosphoglycerate Mutase – Reaction 8

Similar mechanism tophosphoglycerate mutase(glycolysis)- different catalytic residue


Complication! – The Liver

Glycogen is primarily stored in the liver and is used to maintain bloodglucose levels between meals

But … neither G1P nor G6P can be transported out of liver cells

Require separate pathway (below) to convert G6P to glucose for transport


Dietary Polysaccharides

Dextrin + n H20 → n D-glucose Dextrinase

Maltose + H20 → 2 D-glucose Maltase

Lactose + H20 → D-galactose + D-glucose Lactase

Sucrose + H20 → D-fructose + D-glucose Sucrase

Di- and polysaccharides are converted to monosaccharides, then funneled into the glycolytic sequence


Fructose entry into Glycolysis

Two routes for fructose entryinto glycolysis - tissue specific


Fructose entry into Glycolysis

Non-LiverD-Fructose is phosphorylated by hexokinase and F6P enters glycolysis:

Fructose + ATP → fructose 6-phosphate + ADPMg2+

Liver D-Fructose phosphorylated by fructokinase (at C1):

Fructose + ATP → fructose 1-phosphate + ADPMg2+

Fructose 1-phosphate is then cleaved to glyceraldehyde and dihydroxyacetone phosphate (DHAP) by fructose 1-phosphate aldolase.

Glyceraldehyde is phosphorylated by triose kinase and ATP to glyceraldehyde-3-phosphate.DHAP and glyceraldehyde-3-phosphate

Are both glycolytic intermediates


Galactose entry into the Glycolysis

Galactose entry into glycolysis ismore complex than for other dietarysugars


Galactose conversion to Glucose-1-phosphate

Metabolism of Galactose involvesthree enzymes and a sugar nucleotide.

C1 carbon is activatedas phosphate ester

Textbook (3rd Edition) has typo that is corrected here

Glycolysis


Galactose conversion to glucose-1-phosphate

Activation of C1 phosphatevia formation of phosphateester with UDP

→ glycolysis

UDP-glucose + Galactose-1-phosphate↓

UDP-galactose + Glucose-1-phosphate

Must regenerate UDP-glucose tocontinue cycle


Conversion of UDP-galactose to UDP-glucose

Textbook (3rd Edition) has typo that is corrected here

iii. metabolism glucose catabolism – part...

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