Gluconeogenesis

• Gluconeogenesis is a major regulatory process in the liver and kidneys

by which noncarbohydrate substrates; namely glycerol, lactate,
propionate, and glucogenic amino acids; are converted to glucose 6-
phosphate (Glc-6-P), and then to either free glucose or glycogen.
• The liver and kidneys are the major organs containing the full
complement of gluconeogenic enzymes:
• Pyruvate carboxylase
• PEP carboxykinase
• Fructose 1,6-bisphosphatase
• Glucose 6-phosphatase
• Gluconeogenesis is needed to meet the demands for plasma glucose
between meals, which then becomes particularly important as an
energy substrate for nerves, erythrocytes, and other largely anaerobic
cell types.

• Failure of this process can lead to brain dysfunction, coma, and death.
In ruminant animals and carnivores, hepatic gluconeogenesis is a
continual, ongoing process that has little or no relation to the
frequency of food consumption.
• gluconeogenic mechanisms in the liver are used to clear certain
products of metabolism from blood
• (e.g., lactate, produced by erythrocytes, the retina, kidney medulla,
and anaerobic muscle fibers
• glycerol which is continuously produced by adipose tissue when fat is
being mobilized for energy purposes)
• Of the amino acids transported from muscle to liver during starvation,
alanine (Ala) predominates
• The majority of amino acids form TCA intermediates and pyruvate,
and are therefore glucogenic
• Others form acetyl-CoA and, thus, are ketogenic. Acetyl-CoA is not a
(net) substrate for gluconeogenesis since its two carbon atoms are
lost as CO2 in the TCA cycle. Especially important for gluconeogenesis
are Ala (in the liver), and glutamine (in the kidneys)
• It has been estimated that the contribution of amino acids to
ruminant gluconeogenesis is about 5-7% of glucose produced in both
the fed and fasted state.
 Lactate
• A good source of glucose during exercise is lactate (Cori) Cycle), and
during concentrated carbohydrate feeding (where rumen lactate
concentrations are greatly increased), lactate becomes an important
hepatic gluconeogenic substrate. Lactate is also readily oxidized in
cardiac muscle
 Glycerol
• Since glycerol kinase activity is absent in white adipose tissue,
glycerol becomes a waste product of lipolysis, and is converted to
glucose in the liver, and to a lesser extent in the kidney. This
substrate becomes a significant source of glucose in hibernating
animals (e.g., the black bear), where lipolysis becomes necessary for
survival.
 Propionate
• Propionate, a volatile fatty acid (VFA) produced from microbial
carbohydrate digestion in ruminants and other herbivores; is a major
hepatic gluconeogenic substrate.

• The percentage of glucose derived from propionate in the liver varies

with diet (and species), from a maximum of about 70% under heavy
grain feeding in ruminants, to very little during starvation.
• The importance of propionate as a gluconeogenic substrate is
illustrated by the observation that the lactating udder of the goat
may utilize 60-85% of glucose produced by the liver for milk
production.
• In contrast to propionate, acetate and butyrate, the other two major
VFAs produced through microbial carbohydrate digestion, do not
contribute carbon atoms directly to the net synthesis of glucose.
• Certain glucogenic amino acids (namely isoleucine,
valine, threonine, and methionine), the terminal 3 carbons
of odd-chain fatty acids undergoing mitochondrial β-oxidation, and
the β-aminoisobutyrate generated from thymine degradation,
can also enter hepatic gluconeogenesis at the level of propionyl-CoA.
• Entry of propionate into gluconeogenesis (as well as amino acids that
are converted to propionyl-CoA), requires pantothenate (a
source of coenzyme A.SH), vitamin B12, and biotin. These
vitamins are normally synthesized by microbes inhabiting the
digestive tract
 Gluconeogenic Enzymes
• Pyruvate Carboxylase
• The metabolic route from pyruvate to OAA and then on to PEP is
called the dicarboxylic acid (DCA) shuttle. It allows pyruvate and
compounds that can be converted to pyruvate, such as lactate and
amino acids, to be metabolized to PEP without having to traverse the
physiologically irreversible step from PEP to pyruvate in the opposite
direction. Pyruvate carboxylase is a mitochondrial
enzyme that is activated by acetyl-CoA
 PEP Carboxykinase
• Cytoplasmic conversion of OAA to PEP is catalyzed by PEP
carboxykinase, which requires GTP to drive this reaction energetically
in an uphill direction.
• This high energy compound (GTP) is normally derived from the
conversion of succinyl-CoA to succinate in the TCA cycle.
• In most mammals the PEP carboxykinase enzyme is located
predominantly in the cytosol. However, in the guineapig liver, 50% of
PEP carboxykinase activity is found in mitochondria, and in birds this
figure approaches 100%.
• This reaction is the focal point of gluconeogenesis, and the rate-
limiting step. PEP carboxykinase, like fructose 1,6-bisphosphatase and
glucose 6-phosphatase below, is stimulated by the diabetogenic
hormones (i.e., epinephrine, growth hormone, glucagon, and the
glucocorticoids), and inhibited by insulin. Thyroxine also plays a
stimulatory role.
• Epinephrine and glucagon act by increasing intracellular cyclicAMP
levels, and the glucocorticoids act by increasing the synthesis and
activity of the four major gluconeogenic enzymes.
• Although gluconeogenesis also takes place in proximal renal tubular
epithelial cells of the kidney, neither insulin nor glucagon are thought
to affect that process.
 Fructose 1,6-Bisphosphatase
• This enzyme is inducible, and in addition to the diabetogenic
hormones above, is activated by high cytoplasmic ATP and citrate
levels.
• This reaction bypasses the regulatory reaction of glycolysis catalyzed
by phosphofructokinase (PFK), which is inhibited by glucagon, ATP,
phosphocreatine, H+ and citrate
 Glucose 6-Phosphatase
• In liver and kidney tissue (but not in muscle), this enzyme is present
to remove phosphate from Glc-6-P, enabling glucose to diffuse into
blood.

• This is usually the final step in gluconeogenesis and in hepatic

glycogenolysis, which is reflected by a rise in the blood glucose
concentration. When hepatic Glc-6-P is being overproduced through
gluconeogenesis (e.g., during glucocorticoid stimulation), then some
may pass on into glycogen formation, thus replenishing intracellular
reserves
 Glycerol Kinase
• In addition to the four enzymes above, glycerol kinase, with the
assistance of ATP, converts glycerol (from white adipose tissue) to
glycerol 3-P

• It is also a gluconeogenic enzyme and is found in the liver, kidneys,

intestine, brown adipose tissue, and the lactating mammary gland.
• In summary, gluconeogenesis is the process whereby
noncarbohydrate substrates such as glycerol, lactate,
pyruvate, propionate, and gluconeogenic amino
acids are converted to either free glucose or glycogen.
 Glucose transporters (GLUTs)
• Entry of glucose into cells is thought to be carrier-mediated by
integral membrane glucose transporters (GLUTs), that help to move
monosaccharides down its concentration gradient

• Two Na+ -dependent glucose transporter (SGLT) isoforms, and five

GLUT transporter isoforms have been identified in various tissues
• The SGLT 1 and 2 transporters are found in luminal (apical)
membranes of proximal renal tubular epithelial cells

• While both the SGLT 1 and GLUT 5 transporters are found in luminal
(apical) membranes of mucosal cells in the small intestine
• The SGLT 1 transporter requires 2 Na+ /glucose molecule, while the
SGLT 2 transporter requires only 1, and both are insulin-independent.

• The SGLT 1 and/or 2 transporters are responsible for secondary active

transport (i.e., symport) of glucose (and galactose) out of the
intestine and renal tubular filtrate, against their concentration
gradients, and therefore are associated with aerobic processes (i.e.,
generation of ATP for Na+ /K+ -ATPase utilization).
 GLUT 1
• GLUT 1 is responsible for glucose reabsorption across the basolateral
membrane of proximal renal tubular epithelial cells (S-3 segment),
and also glucose uptake into erythrocytes, the colon, and across the
placenta, and blood-brain barrier, e.t.c.
 GLUT 2
• GLUT 2 is the pancreatic insulin-secreting b-cell sensor, and also
functions in transporting glucose out of intestinal and proximal renal
tubular epithelial cells, and also into and out of the liver
 GLUT 3
• GLUT 3 is responsible for basal glucose uptake in neurons, the
placenta, and other organs.
• GLUT 3 and GLUT 1, respectively, may also be involved in the
transport of dehydroascorbate (DHC) out of neurons and into
astrocytes of the brain.
 GLUT 4
• The GLUT 4 transporter is the only one that has been identified as
being insulin-dependent, and is found primarily on plasma
membranes of muscle and fat tissue.
• Transport activity of GLUT 4 appears to be increased through
activation of protein kinase C (PKC)
• In adipocytes the GLUT 4 transporters are translocated from an
inactive pool in the Golgi apparatus to active sites on the plasma
membrane following the binding of insulin to its receptors.

• Following the diffusion of glucose into the cell and the removal of
insulin from its binding sites on plasma membrane receptors, the
GLUT 4 transporters are translocated back to the Golgi.
 GLUT 5
• GLUT 5 transporter is associated with lesser amounts of anaerobic
glucose and fructose (facilitated) diffusion into duodenal and jejunal
mucosal cells.
• Once inside the cell, glucose is rapidly phosphorylated to glucose-6-
phosphate (Glc-6- P), a "trapping" reaction that serves to keep this
important hexose in the cell (since the charged phosphate group
precludes diffusion of glucose back out of the cell).
• The Glc-6-P thus formed now becomes a substrate from which
multiple pathways branch out. It may be used in the Embden-
Meyerhoff pathway (EMP), in glycogen synthesis in hepatocytes and
muscle cells, in the hexose monophosphate shunt (HMS), and in the
uronic acid pathway of hepatocytes.
• The uronic acid pathway is another cytoplasmic route for the
metabolism of Glc-6-P. UDP-Glucuronate, formed through the uronic
acid pathway, is used for numerous hepatic conjugation reactions.
• Cats have difficulty conjugating, and therefore excreting various drugs
from the body that are typically metabolized through the hepatic
glucuronide conjugation mechanism.
• Vitamin C is synthesized via the uronic acid pathway in most animals
except primates, fish, flying mammals, songbirds, and guinea pigs.
 Glycogen Storage Diseases
• Glycogen storage diseases are a group of inherited metabolic
disorders characterized by deficient mobilization of glycogen or
deposition of abnormal forms, leading to muscular weakness,
exercise intolerance, and sometimes death.

• Glycogen storage diseases are known to exist in dogs (usually

miniature breed puppies), cats, horses, and primates, and are
generally characterized by an inability to form or degrade glycogen in
normal metabolic pathways.
• Examples are a type I von Gierke's like disease (a Glc-6-Pase
deficiency), and a type II Pompe's-like disease (where glycogen
accumulation is minimal due to a glycogen branching enzyme
deficiency (GBED)).

• Glycogen branching enzyme belongs to the α-amylase family of

enzymes, which include α-amylases, pullulanas/isoamylase,
cyclodextrin glucanotransferase (CGT).
• Clinical signs in type II glycogen storage disease are apparently related
to cardiac and skeletal muscle glycogenosis, and include gastric reflux
and megaesophagus, systemic muscle weakness, and cardiac
abnormalities

• A type III Cori's-like disease also exists in dogs, which is a debranching

enzyme deficiency.
• Examples of debranching enzymes include 4-α-D-glucanotransferase or
glucosyltransferase and Amylo-α-1,6-glucosidase or glucosidase
• As indicated above, these diseases may result in either the inability to
form glycogen, or in glycogen accumulation in the liver, muscles,
kidneys, and nervous tissue.

• Affected patients become exercise intolerant, blood glucose levels are

low, and hyperlipidemia, hepatomegaly, and ketonemia develop.
• Another glycogen storage abnormality is exhibited in dogs with steroid
(i.e. glycocorticoid) hepatopathy. Affected animals develop
hepatomegaly with excessive glycogen accumulation (vacuolar
hepatopathy).
 The Blood Group
• The human ABO blood groups illustrate the effects of
glycosyltransferases.
• Carbohydrates are attached to glycoproteins and glycolipids on the
surfaces of red blood cells. For one type of blood group, one of the
three different structures, termed A, B, and O, may be present (Figure
• These structures have in common an oligosaccharide foundation
called the O (or sometimes H) antigen.
• The A and B antigens differ from the O antigen by the addition of one
extra monosaccharide, either N-acetylgalactosamine (for A) or
galactose (for B) through an -1,3 linkage to a galactose moiety of the
O antigen.
• Specific glycosyltransferases add the extra monosaccharide to the O
antigen. Each person inherits the gene for one glycosyltransferase of
this type from each parent.
• The type A transferase specifically adds N-
acetylgalactosamine, whereas the type B transferase
adds galactose.
• The O phenotype is the result of a mutation that leads to premature
termination of translation and, hence, to the production of no active
glycosyltransferase.
The Blood Group