carbohydrate metabolism 1
TRANSCRIPT
-
8/8/2019 Carbohydrate Metabolism 1
1/9
LEGEND :
= Clinical correlation;
= Step number (Glycolysis)
= Positive regulation
= Negative regulation
= increase/d
=decrease/d
Glucose : C 6 H 12 O 6 Beginning or end of major pathways of carbohydrate
metabolism Major form in which carbohydrates absorbed form GIT is
presented to cells Major fuel for the brain Biomedical importance
o Provide ATP in absence of oxygen allowing tissues to survive anoxic episodes
o Heart muscle is adapted to aerobic performance glycolytic pathway; poor survival under
conditions of ischemia or myocardialinfarction
hemolytic anemia: pyruvate kinase deficiencyfatigue: phosphofructokinase deficiencycancer cachexia: lactate; gluconeogenesis;
hypermetabolism lactic acidosis: impaired activity of pyruvate
dehydrogenaseRela t ionsh ip of Glucose to Major pa thways of
Carbohydra te Metabol i sm
GLYCOLYSIS (Harpers Illustrated Biochemistry Chapter 18) A.k.a. Embden-Meyerhoff Pa thway A Catabolic Pathway (TCA = Amphibolic Pathway) Used by all cells to extract energy from glucose Aerobic or Anaerobic
A) Aerobic glycolysis Pyruvate will be oxidized to CO 2 ,H2 O, and Energy (ATP);
B Anaerobic glycolysis
3 S tages of Glycolys i s
I. Pr imary S tage
D-Glucose + 2ATP D-fructose 1,6Bisphosphate +2ADP + 2H
A.k.a. Trapping stage trapping of glucose in formof Glucose 6-Phosphate (G6P) with the utilizationof ATP. Phosphorylation with a negatively charged
PO 4 to Glucose forming G6P, glucose is preventedfrom moving outside of the cell therefore trapping it inside the cell.
In von Gierke disease , aka Type I Glycogensosis(Glycogen storage disease) deficiency of Glucose-6-Phosphatase (G6Pase) which catalyzes thehydrolysis of G6P to D-Glucose, preventing G6P tobe converted to D-glucose which can be
transported outside the cell. This results in theaccumulation in the liver of excessive amounts of normal glycogen.
II. Spl i t t ing S tage
D-fructose 1 ,6-Bisphosphate 2 D-Glycera ldehyde 3-Phosphate
Splitting of FBP into two Tr iose phosphates(G3P)
III. Oxido- reduc tase Phosphory la t ion S tage
2D-G3P + 4ADP + 2Pi + 2H + 2Lacta te + 4ATP +2H 2 O
Earning stage formation of ATP
* Stage I and II are the investment stage (Investmentof 2 ATPs).
SUBJECT: BIOCHEMISTRY
TOPIC: CARBOHYDRATEMETABOLISM 1 (GLYCOLYSIS &GLUCONEOGENESIS)
LECTURER: DR. UY
DATE: DECEMBER 2010
#
-
8/8/2019 Carbohydrate Metabolism 1
2/9
GLYCOLYSISSTEP 1 - Phosphory la t ion by
Hexokinase /Glucokinase ( I r revers ib le )- Trapping of glucose by phosphorylation to G6P
DIFFERENCE BETWEEN HEXOKINASE AN DGLUCOKINASE
*This is wherethe 1 st ATP is invested.
Hexokinase :- Low Km for glucose relative to its concentration in
blood. (high affinity for glucose)- Abundant in the Muscles
o Hexokinase immediately reacts even with alow concentration of glucose.
o This rapid reaction of the enzyme isimportant for G6P to be readily available in
the muscles.- Strongly inhib i ted by its product Glucose 6-
Phosha te (G6P) hence its reaction is not atequilibrium.o When there is enough G6P in the muscle
Hexokinase is inhibited. (Low Vmax)- In liver, hexokinase is saturated under normal
conditionGlucokinase (GK) :
- S 0.5 for glucose is higher than the Km for glucose of other kinases
- less sens i t ive to product inhibition by G6P(G6P does not inhibit Glucokinase )
- contribute to the capacity of the liver to bufferblood glucose levels because of the following
GLUT4 (in the muscles and adipose tissues)insulin dependent transporters
In DM type 1 patients, lack of insulin causesfatigue due to lack of ATP caused by thelack of glucose transported inside.
GLUT2 (in liver and pancreas)
GKRP attaches to glucokinase in its inactive form in thenucleus forming a complex with GK.
With the increase in glucose concentration in the cell,release of GK by GKRP into the cytosol is promoted converting GK to its active form. While the increase of Fructose 6-Phosphate (F6P) signals the inactivation of GKback into the nucleus with GKRP therefore inhibiting itsactivity.
M ATURITY-ONSET D IABETES OF THE YOUNG (MODY)Type 2
An autosomal dominant disorder involving mutations in the glucokinase (GCK) gene.
Patients have progressive hyperglycemia that is usuallyasymptomatic at diagnosis and is usually managedwi th d ie t a lone .
STEP 2 I somer isa t ion by Phosphoglucoseisomerise (Reversible)
1
2
Stage I and !! :Investmentphase
Stage III : Energygeneration phase
-
8/8/2019 Carbohydrate Metabolism 1
3/9
-
8/8/2019 Carbohydrate Metabolism 1
4/9
ALDOLASE A ( found in RBCs and musc le )
- Absence of the enzyme (aldolase) may causenonspherocytic hemolytic anemia.
*most/almost all of glycolytic enzyme deficienciesmanifest hemolytic anemia because this causes thepumps in the membrane of RBC do not functionproperly. This allows the RBC to be exposed tooxidative properties which causes hemolysis or
swelling of the RBC.
*In this disease however the hemolytic anemia isnonspherocytic which means the RBCs are not
spherical or sphere-shaped like most hemolyticanemia.
TRIOSE PHOSPHATE ISOMERASE (TPI)
- Patients with TPI deficiency have neonatal hemolyticanemia and progressive neurologic involvement
- Progressive hypotonia with eventual diaphragmparalysis and cardiomyopathy.
STEP 5 Most DHAP are conver ted to G3Pwhich i s u t i l i zed in the g lycoly t ic pa thway.
*because most DHAP is converted to G3P there should be2 molecules of G3P converted to Pyruvate.
STEP 6 ox ida t ion by G3P dehydrogenase
As previously mentioned most of the DHAP will beconverted to G3P, which G3P is the one utilized in theformation of Pyruvate, therefore from everything beginning in step 5 happens in pair.
In this step, G3P is converted to a highenergy compound 1 ,3 -Bisphosphoglycera te (1 ,3BPG) by Glycera ldehyde 3-phosphate dehydrogenase(G3PDH or GAPDH) the reduction of NAD+ to NADH+H +which will form (3)ATP* in the ETC or utilized by the
pyruva te lac ta te lac ta te
dehydrogenase
9
10
4
5
x 26
x 2
7
8
-
8/8/2019 Carbohydrate Metabolism 1
5/9
STEP 7 1 st subs t ra te leve l Phosphory la t ion byPhosphoglycera te k inase (PGK) (and ADP ATP)and format ion of 2 ,3BPG byproduct .
At this step,
1,3BPG is converted to 3-Phosphoglycera te by transferring of the phosphate to ADP forming ATP. This is the first substrate level phosphorylation in Glycolysis. 2ATPsshould be formed because there are 2 molecules of 1,3BPG from the splitting. At this point of glycolysis the2ATPs invested in steps 1 and 3 has already been regained.
There is also the presence of an enzyme mutase ,abundant in the RBC, which may form a byproduct 2,3BPG from 1,3BPG . 2,3BPG shifts the oxygen dissociation curveto the right releasing more oxygen to the tissues. 2,3BPGcan then be converted to 3-Phosphoglycera te (3PG) with the use of the enzyme Phosphatase .
*2,3BPG or diphosphyglycerate/DPG is also likely anintermediate of the reaction.
* However, this pathway does not produce ATP.
*PGK is inhibited by arsenate
Arsenic tox ic i ty arsenate competes with theinorganic phosphate (P i) hydrolyzing 3PG to 2PGwithout ATP formation
STEP 8 I somer iza t ion by Phosphoglycera temutase
3-Phosphoglycerate is isomerized to 2-Phosphoglycerate
STEP 9 Dehydra t ion ca ta lyzed by Enolase
2-Phosphoglycera te removes H 2 O to form a high energy phosphate compound phosphoenol pyruva te (PEP)
*enol compounds=high energy compounds
Enolase is inhibited by Flouride ; used to inhibitglycolysis in blood samples used for measuring glucose.
STEP 10 2 nd Subs t ra te leve l phosphory la t ion(Irreversible)
At this step, phosphate is transferred from PEP to ADP bypyruva te k inase(PK) forming 2 molecules of ATP perglucose. Since the invested ATPs were already regained,
this 2 nd substrate level phosphorylation is the first gaining of ATP from glucose. PK is allosterically activated byFBP (product of the rate limiting step page2)
Pyruvate Kinase Def ic iency & Hemoly t icAnemia
Lack of ATP affects ion pumps especially Na + /K + ATPase. causing the cells to swell & lyse
During starvation , g lucagon is increased. This stimulates the activity of adenyly l cyc lase which converts ATP tocAMP. cAMP then activates PKA. PKA phosphorylatesPyruvate kinase(PK) turning it into its inactive form.Therefore, with decreased glucose glycolysis decreaseproduction of pyruvate with the inactivation of PK.
*kinases are inactive when phosphorylated similar to PFK-2
During Anaerobic glycolysis pyruva te forms Lacta tecatalyzed by the enzyme Lacta te dehydrogenaseutilizing with it. NADH+H + from the activity of G3Pdehydrogenase
2 ATP consumpt ion with theenzymesHexokinase/Glucokinase andPhosphofructoki nase-1 (PFK-1).
2NADH+2H + p roduced fromG3P dehydrogenase .
4ATPproduced from(2) Phospho-
glyceratekinase (PGK)and (2) (PK Pyruvatekinase )
In anaerobicpathway, 2NADH
-
8/8/2019 Carbohydrate Metabolism 1
6/9
3 IRREVERSIBLE STEPS OF GLYCOLYSIS WHICH ARE ALSOREGULATORY STEPS
*All are kinase enzymes.
NADH Generated by Glycolysis has to be oxidized back toNAD+. This is done by the enzyme LactateDehydrogenase(anaerobic) & Substrate Shuttles(aerobic).
In Aerobic glycolysis, NADH produced by G3PDH needs tobe transported into the mitochondria for ATP synthesis in the ETC, however, NADH is NOT permeable to the innermitochondrial membrane. By utilizing substrates (Malateand Aspartate) permeable to the inner mitochondrialmembrane, NADH is made available to the ETC in themitochondria.
OAA + NADH Malate Dehydrogenase Malate + NAD+
Malatedehydrogenase (MDH) Oxaloace ta te OAA Mala te
which is also permeable to the inner mitochondrialmembrane then exits the mitochondria and converted back
to OAA by AST.
Fate of the PYRUVATE
Carboxylation of Pyruvate toOxaloace ta te (OAA) iscatalyzed by Pyruvatecarboxylase .
(Oxidative) Decarboxylation of Pyruvate to Acety l CoA iscatalyzed be Pyruvate deydrogenase complex along with
the reduction of NAD +. Acetyl CoA will proceed to TCA cycle .The Pyruvate dehydrogenase complex is a multienzymecomplex which consists of Pyruvate Dehydrogenase,Dihydrolypoyl Tranacetylase, Dihydrolipoyl Dehydrogenase.
(The succeeding discussion on Oxidative decarboxylation was notdiscussed but it was in the PPT and is also found in the Harperson chapter 18, page 153, The oxidation of pyruvate to Acetyl-CoAis the irreversible route from glycolysis to the citric acid cycle.)
During the conversion the intermediates do not dissociate
and remain bound to the components of the multienzymecomplex.
MECHANISM: (OXIDATIVE CARBOXYLATIONOF PYRUVATE)
Pyruvate is decarboxylated to a hydroxyethyl (Acyl) bound to thiamine diphosphate (TPP or TDP) with the enzrymePyruvate dehydrogenase, forming Acyl-TPP. Acyl-TPP reactswith an oxidized lipoamide (Lip-S 2 ) catalyzed be theDihydrolipoyl Transacetylase component, forming Acyl-Lipoate/Acetyl lipoamide (in Harpers). Acyl-lipoate will thenreacts with Coenzyme A(CoA-SH) forming Acetyl CoA and
the reduced lipoamide. However lipoamide has to bereturned to its original form which is reoxidized by theDihydrolipoyl Dehydrogenase(flavoprotein componentcontaining an FAD, forming FADH and Lip-S 2 . DihydrolipoylDehydrogenase is finally oxidized by NAD + for it to return toits orginal form flavoprotein containing FAD.
-
8/8/2019 Carbohydrate Metabolism 1
7/9
-
8/8/2019 Carbohydrate Metabolism 1
8/9
2Lactate + 6ATP + 6H 2 O Glucose + 6ADP + 6P i + 4H +
Glucose can be immediately trapped by the muscle with theenzyme Hexokinase. During exercise there will be anincreased amount of lactate causing lact ic acidosis . Thiswill cause pain in the muscles (commonly in the form of cramps). Because of muscles do not have lac ta tedehydrogenase Lactate will be transported to the liverwhere it can be converted to glucose. In the liver lac ta tedehydrogenase can catalyze the oxidation of Lac ta te to
Pyruva te which requires 6ATP to form glucose(gluconeogenesis) to be sent back to the muscle and other
tissues. Utilizing 6ATP makes gluconeogenesis an expensivepathway.
Amino ac ids as subs t ra te for Gluconeogenes is
In the muscles Alanine t ransaminase or Alanineaminotransferase (ALT) catalyze the transferring of anamino group from glu tamate to pyruva te forming -ke toglu ta ra te and a lan ine . Through the blood Alanine
travels to the liver where it is converted back to pyruvatewith ALT. Aside from the 6 ATP required for pyruvate toundergo gluconeogenesis, Ammonium needs to bereleased from glutamate through urea cycle utilizing 4 ATP.This makes amino acids as substrate more expensive (ATP).
2Ala + 10ATP + CO 2 Glucose + Urea + 10ADP + 10P i
Excessive amounts of ammonia loaded into the liver, up to a point that not everything can be immediatelyconverted to urea can cause comatose due to hepaticencephalopathy. This is common in liver cirrhosiscaused by alcoholism.
GLUCOKINASE IN THE LIVER Regulates blood glucose after a meal Promotes increased hepatic utilization of glucose
o Promotes Glycolysis not gluconeogenesisnor glycogenolysis.
Carboxyla t ion of Pyruva te is catalyzed by Pyruvatecarboxylase by transferring the CO 2 attached to biotinforming Oxaloace ta te (OAA).
Because OAA is not permeable to the mitochondrialmembrane it is reduced by NADH to mala te .
In the cytosol, mala te is reoxidized to OAA with NAD +.
OAA will then be converted (decarboxylation andphosphorylation) to Phosphoenol pyruva te (PEP ) byPEP carboxykinase (PEPCK) with the cofactor GTP( GDP) .
The succeeding steps beginning from PEP are reversereactions of glycolysis,except in FBPase &G6Pase.
ATP is again utilizedin the formation of 1,3-BPG.
Because 2 moleculesis required in theformation of FBP 2pyruvate must beutilized to form 1molecule of glucose.
After this step, 4 ATPand 2GTP has beenutilized by the pathway.
2NADH will beutilized to convert 2moleculesof1,3BPG to form 2molecules of G3P.
PFK-1 is anirreversible enzymeof glycolysis
thereforegluconeogenesisneeds its ownenzyme FBPase.
The last step of gluconeogenesis is catalyzed by Glucose-6-phosphatase(G6Pase) which dephosphorylates G6P
to Glucose . G6P is not found in the muscles therefore,gluconeogenesis cannot occur in the muscle
von Gierkes Disease (mentioned in page 1 ) - type I glycogen storage disease(GSD)- deficiency of glucose-6-phosphatase- severe hypoglycemia causing lethargy,seizures and
brain damage- hepatomegaly, increased bleeding (clotting factors
2,6,7,9,10 and platelets are affected) and growthretardation
Glucagon Maingluconeogenic
hormone
Similar to theprevious discussionin page 3 and 5,
-
8/8/2019 Carbohydrate Metabolism 1
9/9
Catecholamine increase secretion of Glucagon which issynthesized from -pancreatic cells.
G LUCOSE IS S YNTHESIZED FROM M OS T AMINO ACIDS All amino acids except leucine & lysine can supply
carbon for the net synthesis of glucose by
gluconeogenesis Pyruvate, oxaloacetate catabolic products of amino
acids from which glucose synthesis can occur Anaplerotic Reactions those that lead to net
synthesis of TCA intermediates
Hepat ic encephalopa thy utilization of aminoacids for gluconeogenesis may cause production of excess urea which may lead to an encephalopathyassociated with cirrhosis of the liver, attributed to thepassage of toxic nitrogenous substances from theportal to the systemic circulation; cerebralmanifestations may include coma.
Gluconeogenic Ketogenic Both
GlycineSerineValineHistidineArginineCysteineProlineHydrodyprolineAlanineGlutamateGlutamineAspartateAsparaginesMethionine
LeucineLysine
ThreonineIsoleucinePhenylalanineTyrosineTryptophan
Glucose can be synthes ized f rom Propionyl -CoA(odd-numbered carbon) Propionyl-CoA good precursor for gluconeogenesis as
it yields oxaloacetate by anaplerotic pathway(TCA cycle). Triacylglycerol(TAG) when hydrolyzed yields 3 FAs &
glycerol (a substrate for gluconeogenesis)
o Because Glycerol kinase(found in adipose tissue) is absent in the liver, TAG needs to be hydrolyzed
During starvation a great amount of ATP is used forgluconeogenesis which may be taken from the oxidation of fatty acids in the liver. This process however forms ketonebodies.Insu l in
the only counter-regulatory hormone in gluconeogenesis main glycolytic hormone action
o binds to receptoro receptor sends signal activating phosphoprotein
phosphataseo inactive PFK-2 becomes active PFK-2 (Pi removed)o active PFK-2 increases concentration of F-2,6 BPo F-2,6 BP binds to PFK-1 & increases its activityo Rate of glycolysis is increased
Alcohol Oxida t ion Inh ib i t s Gluconeogenes is
Due to excess NADH forcing the equilibrium of thereactions catalyzed by lactate dehydrogenase and malatedehydrogenase
PEPCK Deficiency- rare but severe metabolic defect
- absence of the cytosolic form results in cerebra la t rophy , opt ic a t rophy , fatty infiltration of the liverand kidney and intractable hypoglycemia (including lost of vision)
Fruc tose-1 ,6-Bisphosphatase Def ic iency
- Presents with neonatal hypoglycemia, along withacidosis, irritability, tachycardia, dyspnea, hypotonia,moderate hepatomegaly.
- typically only the liver enzyme is deficient, the muscleactivity is normal
(Please refer to slide 82 of the PPT for the gluconeogenesis summary)