CARBOHYDRATE

METABOLISM I
Dr. Jonathan Lim Chee Woei
(cheewoei@upm.edu.my)
Pharmacotherapeutics Unit
Department of Medicine
FMHS
 Glycolysis
 Aerobic VS Anaerobic Glycolysis
 Glycolytic pathway
- Reaction sequences and enzymes
- Priming stage or phosphorylation of glucose, splitting stage and oxidoreduction stage and
synthesis of ATP
- Control mechanism of glycolysis
- Key regulatory enzymes - glucokinase, phosphofructokinase and pyruvate kinase
 Alternate fates of pyruvate
- Lactate metabolism and clinical correlations: lactic acidosis

 Gluconeogenesis
 Importance of producing blood glucose
 Pathways of gluconeogenesis-from lactate, pyruvate, glucogenic amino acids
 Energy cost of gluconeogenesis
 Reciprocal regulation of glycolysis and gluconeogenesis
 Alanine and cori cycle

 Regulation of carbohydrate metabolism by insulin, glucagon and

epinephrine

 Clinical correlations- lactic acidosis, hyperglycaemia & diabetic mellitus;

pyruvate kinase deficiency
Major pathways of carbohydrate metabolism.
Glycolysis
What is glycolysis?

• Glycolysis is central pathway in generating energy

and metabolic intermediaries.

• All carbohydrates to be catabolized must enter the

glycolytic pathway

• Ten step metabolic pathway to convert glucose into

2 pyruvate + 2 NADH + 2 ATP

• Occurs in cytoplasm.

• Does not require oxygen

Aerobic (Blue) VS Anaerobic glycolysis (Green)
Glycolysis

Glucose
hexokinase ATP
ADP

Glucose-6-phosphate
phosphoglucoisomerase

Fructose-6-phosphate
ATP
phosphofructokinase ADP

aldolase Fructose-1,6-bisphosphate

Triose phosphate isomerase

Glyceraldehyde 3-phosphate Dihydroxyacetone phosphate
Cont’d

Glyceraldehyde 3-phosphate
Glyceraldehyde-3-phosphate deHase NAD+ + Pi
NADH + H+
Glycerate-1,3-Bisphosphate
ADP
Phospoglycerate kinase ATP

Glycerate-3-Phosphate
Phospoglyceromutase

Glycerate-2-Phosphate
Enolase H2O
Phosphoenolpyruvate
ADP
Pyruvate kinase ATP

Pyruvate
Three irreversible kinase reactions primarily drive
glycolysis forward.

 hexokinase or glucokinase

 Phosphofructokinase

 pyruvate kinase
 Key regulatory enzyme

Hexokinase (E.C 2.7.1.1)

 found in all cells of every organism=ubiquitous

 Nonspecific enzyme -Catalyzes the phosphorylation of hexoses such as

glucose, mannose, fructose

 relatively high affinity for glucose, KM = ~0.05- 0.1 mM

2nd substrate is Mg2+-ATP complex
Mg2+ shield α and β of the ATP making γ more accessible for nucleophilic
attach by C6-OH group of glucose

 Can be inhibited by

1. Uncomplexed ATP (competitive inhibitor)

2. Glucose 6-phosphate
 Key regulatory enzyme

Four mammalian isozymes of hexokinase

Types I- housekeeping enzyme unaffected by most

physiological, hormonal, and metabolic change

Types II- muscle and heart; mitochondria outer membrane

Types III- inhibited by glucose at physiologic concentrations

Types IV- Glucokinase is the form of the enzyme found in

hepatocytes  Different
 Key regulatory enzyme
Glucokinase
*in liver

*Primarily involved in maintaining blood glucose

*high KM for glucose; Km is the concentration of substrate which permits the

enzyme to achieve half Vmax

*not inhibited by glucose-6-phosphate

*most effective when glucose level in blood is high, i.e., right after meal.
 Key regulatory enzyme
Glucokinase vs Hexokinase
Hexokinase Glucokinase

Location All tissues Liver & Pancreatic Beta

Cells
Km for glucose ~0.05mM ~5.5mM

Relationship between Hyperbolic Curve Sigmoidal Curve

glucose concentration &
reaction rate
Feedback inhibition by Yes No
Glucose-6-phosphate
Induction by Insulin No Yes
 Key regulatory enzyme

Phosphofructose Kinase
 PFK reaction is similar to hexokinase reaction
 PFK catalyzes the nucleophilic attack by C1-OH grp of F6P on the
electrophilic γ-phosphorus atom of Mg2+-ATP complex
 PFK plays central role in control of glycolysis because it catalyzes one of
pathway’s rate determining reaction
2-
CH2OPO3
2-
O CH2OPO3
2-
CH2OPO3 phosphofructokinase-1
(PFK-1), Mg 2+
O CH OH
2
H HO
H HO H
H
OH OH
OH H OH H
fructose-6-phosphate fructose-1,6-bisphosphate
+ +
ATP ADP
 Key regulatory enzyme
 The committed step! (irreversible) pathway rate determining
reactions
 PFK-1 (E.C 2.7.1.11) is a tetrameric enzyme
 tetramer composed of different combinations of three types of
subunits: muscle (M), liver (L), and platelet (P). The composition of the
PFK1 tetramer differs according to the tissue type it is present in.
 allosteric effectors:
 high ATP conc. depresses rate

 fructose-2,6-bisP activates
 Key regulatory enzyme

Pyruvate Kinase
Glycolysis Reaction 10: Final generation of ATP
O O - rxn is exergonic
C O C O
C O PO3
2-
C O - pyruvate is the primary
Pyruvate kinase
Mg2+ CH3 product of glycolysis
CH2
PEP pyruvate
+ + - pyruvate kinase is a
ADP ATP highly regulated enzyme.
 Pyruvate kinase deficiency
 Severity is highly variable with some have life threatening

 2nd most common genetic deficiency that causes hemolytic anemia (G6PDH
deficiency  Most common)

 PK catalyses conversion of PEP to pyruvate with ATP generation

 Why hemolytic anemia is seen in PK Def?

 Normal RBC has no mitochondria thus depends on glycolysis for ATP

 With PK def lead to reduced rate of glycolysis.

 Lack ATP (lack of energy to maintain rbc structure)
premature death

Blood film shows prickle cells

 Key regulatory enzyme

Enzyme Activator Inhibitor

Hexokinase Glucose-6-
Phosphate, ATP
PFK-1 Fructose-2,6-bis Citrate, ATP
phosphate, AMP
Pyruvate Fructose-1,6-bis Acetyl-CoA, ATP
kinase phosphate, AMP

Allosteric activators such as AMP and ADP bind to the allosteric site as to
facilitate the formation of the R state by inducing structural changes in the
enzyme.

Similarly, inhibitors such as ATP and PEP bind to the same allosteric site and
facilitate the formation of the T state, thereby inhibiting enzyme activity.
Diverse fates of pyruvate

To citric acid cycle

Anaerobic Yeast
Pyruvate  Lactate
• Under anaerobic conditions (red blood cells, parts
of the retina, and in skeletal muscle cells during
strenuous exercise)

• occurs in the cytosol with the help of lactate

dehydrogenase (LDH)

• The activities of glyceraldehyde 3-phosphate DH

and LDH are linked metabolically  Pyruvate is
generated largely by anaerobic glycolysis
(Embden–Meyerhof pathway).

• The blood lactate: pyruvate ratio is normally 10:1,

but it rises with an increased ratio of NADH
concentration ([NADH]) to NAD+ concentration
([NAD+]) (redox state).

• Helps drive glycolysis by using up NADH and

release NAD+  Redox balance is maintained.

Glucose + 2 Pi +2 ADP → 2 lactate + 2 ATP + 2 H2O

Lactate dehydrogenase
 5 isoforms (Combinations of two subunits, LDH-M (encoded by LDH-A) and
LDH-H (encoded by LDH-B))
1. LDH-1 (4H)  heart and RBC
2. LDH-2 (3H1M)  reticuloendothelial system
3. LDH-3 (2H2M)  lungs
4. LDH-4 (1H3M)  kidneys, placenta, pancreas
5. LDH-5 (4M)  liver, striated muscle

 The LDH-M subunit has a higher affinity for pyruvate and its reduction than does LDH-H;
thus, the nature of the LDH isoforms in tissues affects lactate metabolism.

 LDH is general indicator of tissue and cellular damage (Diagnostic

Purpose)
 Detect hemolytic anemia, megaloblastic anemia, severe infections
 Skeletal muscle and liver contain predominantly the “M” forms; heart the “H”
forms. During and after myocardial infarction (heart attack), heart cells die
releasing LDH into the circulation.
 “What happens to the lactate after a run?”

 Lactate is reconverted to pyruvate and consumed in the mitochondria of

the liver, kidney, and other tissues, which have LDH.
Lactate + NAD+ pyruvate + NADH + H+

 The pathways include:

 Cori cycle, which generates glucose but consumes ATP in the liver and kidney
(gluconeogenesis)
 TCA cycle and oxidative phosphorylation in the liver, kidney, muscle, heart, brain,
and other tissues, which generate ATP when pyruvate is oxidized to carbon
dioxide and water.

 Lactate consumption is promoted by intra-organ and inter-organ lactate

shuttles facilitated by monocarboxylic acid transporters (MCTs), which
mediate the influx and efflux of lactate and accompanying protons.

 Normally, the generation and consumption of lactate are equivalent,

which results in a stable concentration of lactate
GLUCONEOGENESIS
GLUCONEOGENESIS
Definition: The biosynthesis of glucose from simpler
molecules, primarily pyruvate and its precursors.

The gluconeogenesis pathway is similar to the reverse of

glycolysis but differs at critical sites.

 control of these opposing pathways is reciprocal so that

physiological conditions favoring one disfavour the other and
vice versa.

General principles of metabolic control

a) pathways are not simple reversals of each other
b) under reciprocal control
Why do we produce glucose?
• Need to maintain glucose levels in a narrow range in blood.
• Some tissues (Brain, Erythrocytes, and Muscles) utilize glucose at a rapid rate
 require glucose in addition to dietary glucose  gluconeogenesis
• The brain and erythrocytes can use only glucose as a source of energy.

• During starvation or periods of limited carbohydrate intake, when the levels of liver
glycogen are low (liver glycogen supplies are adequate for 10-24 hours),
gluconeogenesis is important in maintaining adequate blood sugar concentrations.

• During extended exercise, when CHO and lipid reserves are mobilised,
gluconeogenesis allows the use of lactate from glycolysis and glycerol from fat
breakdown.

• During metabolic acidosis, gluconeogenesis in the kidney allows the excretion of an

increased number of protons.

• Gluconeogenesis allows the use of dietary protein/lipid in CHO pathway.

Where is glucose synthesized?
1. The liver - major location for gluconeogenesis (85%-95% of the glucose that is
made)

2. During starvation or metabolic acidosis, the kidney (Renal Gluconeogenesis) is

capable of making glucose and may contribute up to 50% of the glucose formed
because in these condition, the amount contributed by the liver is decreased
considerably.

3. The only other tissue capable of gluconeogenesis is the epithelial cell of the
small intestine, which contributes not more than 5% of the total glucose
formation.
Precursors/ substrates for gluconeogenesis
 pyruvate- major precursor

 lactate–from muscle, forms pyruvate

 TCA cycle intermediates

 some amino acid carbon skeletons

- from diet or breakdown of muscle protein during starvation
- most important is alanine
- glucogenic amino acids other than alanine may be converted to TCA intermediates that are
metabolised to oxaloacetate, an intermediate in gluconeogenesis.

 propionate
- from breakdown of fatty acids and amino acids.

 Glycerol
- is formed in adipose tissue by lipolysis and dietary triacylglycerols.
- is released into the blood and taken up by the liver where it is phosphorylated to 3-
phosphoglycerate, which is an intermediate in gluconeogenesis.
 Adipose Liver Muscle

Glucose Glucose Glucose

3-phosphoglycerate
TAG

glycerol glycerol pyruvate pyruvate

alanine lactate lactate alanine

Interplay between metabolic pathways in different tissues

 glyceraldehyde-3-phosphate
Summary of
NAD+ + Pi Glyceraldehyde-3-phosphate
Gluconeogenesis NADH + H+ Dehydrogenase
Pathway: 1,3-bisphosphoglycerate
ADP
Phosphoglycerate Kinase
ATP
3-phosphoglycerate
Phosphoglycerate Mutase
Gluconeogenesis
2-phosphoglycerate
enzyme names in Enolase
H2O
red. phosphoenolpyruvate
CO2 + GDP
PEP Carboxykinase
GTP
oxaloacetate
Glycolysis enzyme Pi + ADP
Pyruvate Carboxylase
names in blue. HCO3 + ATP
pyruvate Gluconeogenesis
 glucose Gluconeogenesis
Pi
Glucose-6-phosphatase
H2O
glucose-6-phosphate
Phosphoglucose Isomerase
fructose-6-phosphate
Pi
Fructose-1,6-bisphosphatase
H2O
fructose-1,6-bisphosphate
Aldolase

glyceraldehyde-3-phosphate + dihydroxyacetone-phosphate
Triosephosphate
Isomerase
(continued)
Pyruvate  Oxaloacetate
1. Enzyme: Pyruvate carboxylase (The prosthetic group of the enzyme is biotin. Mg2+ and
Mn2+ is required. Acetyl CoA is required as an activator and regulates activity in a
concentration-dependent manner.)
2. The hydrolysis of ATP provides energy for the carboxylation of pyruvate.
3. Occurs in mitochondrial matrix.
O
O C O
C O Acetyl-CoA C O
Biotin, Mg2+
pyruvate CH2
C O carboxylase
C O
CH3
O
Pyruvate Oxaloacetate
+ +
ATP ADP
+ +
HCO3- Pi
Interconversion of OAA and malate

OAA cannot permeate the mitochondrial

membrane wall, and it must be transported
across the membrane in a form of malate.

• Malate deHase catalyses the reversible

reaction

OAA + NADH + H+ malate + NAD+

(occurs in mitochondria)

In the cytosol, a cytosolic malate deHase

catalyses the reverse reaction, which
generates OAA.
OAA  PEP
1. Hydrolysis of GTP provides the energy for the carboxylation of OAA.
2. PEP carboxykinase (PEPCK) catalyses this reaction, which requires Mn2+
for activation.
3. The two reactions catalysed by pyruvate carboxylase and PEPCK convert
pyruvate to PEP, thus bypassing the irreversible reaction catalysed by
pyruvate kinase during glycolysis.
4. Occurs in cytosol.

PEP  F-1,6-BP
Six sequential conversion steps are catalyzed by the same enzymes of glycolysis.

F-1,6-BP  F-6-P and Pi

1. This reaction is the reverse of the reaction catalysed by
phosphofructokinase during glycolysis.
2. Fructose-1,6-bisphosphatase catalyses this reaction- major regulatory
enzyme in gluconeogenesis.
 Allosteric binder- Citrate as activator; AMP and F-2,6-BP as
inhibitors
F6P to G-6-P
• This reaction, the reverse of the conversion of G6P to F6P in glycolysis, is
catalyzed by the same enzyme, phosphoglucose isomerase.

G6P to glucose and Pi

• This is the reverse of the reaction catalysed by glucokinase and hexokinase
during glycolysis.
• However, in gluconeogenesis this reaction is catalysed by glucose-6-
phosphatase, which is present only in gluconeogenic tissues.
Gluconeogenesis Stoichiometry
2pyruvate + 4ATP + 2GTP + 2NADH + 6H2O

glucose + 4ADP + 2GDP +6Pi +2NAD+ + 2H+

G°’ = -9kcal mol-1.

Just the reverse of glycolysis, G°’ = 20kcal mol-1.

Alanine cycle/ Cahill cycle

 During starvation  muscles degrade amino acids for energy needs, the enzyme alanine
transaminase (muscle, liver, and the intestine) converts L-glutamate and pyruvate into α-
ketoglutarate and L-alanine.

 The resulting L-alanine is released by the muscle into the blood, taken up by the liver, and
converted back to pyruvate by deamination

 The nitrogen enters the urea cycle whereas pyruvate is then used to produce glucose
via gluconeogenesis.
Alanine cycle/ Cahill cycle

• Therefore, this pathway is used instead of the Cori cycle only when an
aminotransferase is present, when there is a need to transfer ammonia to
the liver and when the body is in a state of catabolism (muscle
breakdown).

• Unlike in the Cori cycle, NADH is conserved because lactate is not

formed.
Cori Cycle

 During exercise, pyruvate formed from glucose by glycolysis in muscle and RBC is
converted to lactate by LDH

Lactate released into blood, liver uptake, convert to pyruvate by LDH

Pyruvate converted to glucose via gluconeogenesis in liver

The glucose may travel back to the muscle to fuel glycolysis.

Regulation of carbohydrate
metabolism by insulin,
glucagon and epinephrine
Glycolysis and gluconeogenesis are
reciprocally regulated

Insulin stimulates Glucagon stimulates

A high [AMP] indicates

that the energy charge
is low and signals the
need for ATP.

High [ATP] and

[citrate] indicate the
energy charge is high
and intermediates are
abundant.
epinephrine
Clinical correlations- lactic
acidosis, hyperglycaemia &
diabetic mellitus; pyruvate
kinase deficiency
Lactic acidosis

 Lactic acidosis results from the accumulation of lactate and protons

(low pH) in the body fluids and is often associated with poor clinical
outcomes.

 The effect of lactic acidosis is governed by its severity and the clinical
context.

 Mortality is increased by a factor of nearly three when lactic acidosis

accompanies low-flow states or sepsis, and the higher the lactate level,
the worse the outcome.

 Although hyperlactatemia is often attributed to tissue hypoxia, it can

result from other mechanisms (low blood pressure or blood problems) .

 Control of the triggering conditions is the only effective means of

treatment.
Hyperglycaemia- causes
 Cushing’s syndrome – a collection of hormonal disorders
characterised by high levels of the steroid hormones that act like
cortisol, which is normally produced by the adrenal gland. Causes
include tumours of the pituitary and adrenal glands, certain
tumours in other areas of the body, and steroid drug therapy for
inflammatory disorders
 Pancreatitis – the pancreas makes the hormone insulin.
Pancreatitis is inflammation of the pancreas, which can be either
acute or chronic. Alcoholics are one group at risk of developing
pancreatitis. Other causes of diabetes with chronic pancreatitis
include the inherited conditions cystic fibrosis and
haemochromatosis
 Acromegaly – excess growth hormone secretion
 Certain medications – including some diuretics (drugs that remove
water from the body) and steroids
 Liver disease – such as cirrhosis of the liver.
 Diabetes mellitus
Type 1(DM1) Beta cell destruction-complete lack of insulin
Polyuria, polydipsia, polyphagia, weight loss

Type 2 (DM2) Beta cell dysfunction and insulin resistance

Gestational Beta cell dysfunction and insulin resistance during

pregnancy

others Genetic defects of beta cell function

Exocrine pancreatic diseases
Endocrinopathies
Drug or chemical induced
Other rare forms
 Hypoglycemia & Premature Infants

 Blood glu should be monitored for infants at risk for

hypoglycemia (Decreased glycogen storage)

 Severe hypoglycemia is risk factor for neuronal cell death

and poor neurodevelopmental outcomes

 Neonatal conditions- premature, infection, polycythernia,

hypothermia, perinatal asphyxia
 Hypoglycemia & Alcoholism
 Alcohol inhibits gluconeogenesis

 Hypoglycemia may develop after acute alcohol

consumption

 Alcohol causes hypoglycemia if consumed without

food

 Recovery from acute ethanol poisoning is rapid if liver

and kidneys are healthy