Monday at the Metabolic Clinic
Adventures in Glycolysis
by
Nancy Boury
Iowa State University
1
Background
You have been chosen for a very competitive paid
undergraduate clinical internship position (uCLIP). You
spent your first weeks becoming a certified nurse
assistant (CNA). You will be paid to work as a CNA at St.
Visintainer’s, assisting the nurses on the suspected
metabolic disorder’s acute-care floor.
As part of the internship, you also will be allowed to follow
Dr. Saccharo as he sees clinic patients. Dr. Saccharo is
an expert in enzyme deficiency disorders relating to
glycolysis. Because you hope to one day become a
family physician, you are excited about the opportunity to
learn about these rare metabolic disorders. In order to
prepare you for you first day on the metabolic ward, Dr.
Saccharo requests that you research normal sugar
metabolism, particularly glycolysis.
2
Monday Morning at the Metabolic
Disorders Clinic
After working all weekend on Dr. Saccharo’s
assignment, you feel ready for anything –
including Monday morning. As you ride up
the elevator with your mentor and fellow
interns, Dr. Saccharo asks a few questions
to determine how prepared you are for this
morning’s clinic hours.
3
Please get into your groups of
four and take out your clickers
4
CQ#1: Which of the following enzymes
catalyzes a reaction that produces ATP
during glycolysis?
A. Hexokinase (HK)
B. Phosphoglucose isomerase (PGI)
C. Phosphofructokinase (PFK)
D. Triosephophate isomerase (TPI)
E. Pyruvate Kinase (PK)
5
CQ#2: Which of the following enzymes
breaks one 6-carbon sugar into 2 3-carbon
sugars?
A. Hexokinase (HK)
B. Phosphoglucoisomerase (PGI)
C. Phosphofructokinase (PFK)
D. Aldolase
E. Enolase
6
Dr. Saccharo’s
Review of Glycolysis
Glucose is an important source of chemical
energy, but this energy must be converted
from a large denomination of energy (~680
kilocalories per mole of glucose) to more
usable forms, such as ATP (~7.3
kilocalories per mole). Imagine standing in
front of a vending machine with a $100 bill
vs a $1 bill.
7
Dr. Saccharo’s
Review of Glucose Homeostasis
• Your body (particularly your brain) needs glucose as fuel for
cell processes. When you eat, your blood glucose levels will
temporarily rise, signaling the pancreas to release insulin.
When you have gone without food for several hours, your
pancreas will release glucagon, which triggers the liver to
release glucose from glycogen stores.
• If your blood sugars are too high, your blood vessels are
damaged and the organs they supply with blood are
damaged as well. Consistently high levels of blood sugars
lead to kidney, heart, liver, and brain injury over time.
• If your blood sugars are too low, your brain will starve for
energy. As a result, you could pass out, enter a coma, and
8
potentially die in a matter of hours.
Fed – Blood
Glucose
Levels High
FastingGlucose
Levels Low
Pancreas
Releases
Glucagon
Pancreas
Releases
Insulin
Liver
Glucagon
Receptors
Liver
Releases
Glucose into
the Blood
Body Cells
have Insulin
Receptors
Glucose
Levels
Normal
Body Cells
Take Up
Glucose from
the Blood
9
Dr. Saccharo’s
Review of Glucose Homeostasis
• Diabetics either have cells that don’t respond to
insulin properly (Type II diabetes, or late-onset) or
don’t have functional insulin produced (Type I
diabetes, or juvenile).
• This sugar balance, however, isn’t the only thing
we worry about here in the metabolic disorders
ward. There are genetic disorders where a
person is missing an enzyme required to break
down the glucose (or other sugar) once it gets into
its target cell.
10
• Once glucose enters the cell, hexokinase
will add a phosphate group to the glucose.
This gives the glucose a negative charge
and traps it within the cell.
• This addition of a phosphate group is the
beginning of glycolysis, which is also
known as the Embden Myerhof pathway.
11
Please trade papers within your groups and
check each other’s homework
12
Embden-Myerhof Pathway
13
Clicker Survey: How many mistakes
were in the paper you graded?
A. 0-1
B. 2-3
C. 4-5
D. More than 5
What was the most common mistake
made within your group?
14
Jamie, one of your fellow interns, is bothered
because he knew the clinic frequently treats
people with enzyme deficiency disorders. He
asks Dr. Saccharo:
“If glycolysis is needed to use glucose,
how can a person live without all the
enzymes needed for glycolysis?”
15
One reaction – many different
enzymes
• Isoenzymes use the same reactants to
produce the same products
• They may have:
– Different genes
– Different tissue expression
– Different developmental timing of expression
• They will likely have slightly different
affinities for their substrates.
16
Phosphofructokinase as an example
• Three different forms and three different genes:
– PFK – L is expressed in the liver and the gene for this
isoenzyme is found on chromosome 21
– PFK – M is expressed in the muscle and the gene is
found on chromosome 12
– PFK – P is expressed in platelets and the gene is
found on chromosome 10
• Looking at the homework – this is the same reaction
– Adds a 2nd phosphate to fructose-6-phophate to form
fructose-1,6-bisphosphate
17
CQ#3:If a person would be deficient in all
3 forms of PFK, this person would ___.
A. Have lower than normal glycogen stores.
B. Have more mitochondrial activity than
normal.
C. Be dead.
D. Produce more energy per glucose
molecule because glycolysis would be
unregulated.
18
Time for the clinic to open…
Dr. Saccharo asks the
interns, “Any questions
before we get started
seeing patients?”
Please keep your
corrected homework on
your desk to use as a
reference as we
discuss the patients.
19
Patient #1: Ann is a teenager and avid golfer who was referred
to the clinic after being refused at the blood drive and tiring
easily on the high school golf course during practice.
Levels (+ = normal, - = decreased levels, ++ = increased levels)
Blood Metabolic Panel
Blood Glucose
+
Glucose-6-phosphate
++
Fructose-6-phosphate
++
Fructose-1,6,bisphosphate
++
Glyceraldehyde-3-phosphate
++
1,3-bisphosphoglycerate
++
Phosphoenolpyruvate
++
Pyruvate
-
ATP
-
Red Blood Cell Concentration
-
20
CQ#4: Assuming that Dr. Saccharo is
correctly assuming that Ann has a defect in
glycolysis, what is the most likely defect
based on the blood metabolic profile?
A. Hexokinase
B. Phosphofructokinase
C. Triosephosphate Isomerase
D. Pyruvate kinase
E. Aldolase
21
Patient #2: Marie is a 32-year-old mother of three
complaining of fatigue and muscle cramps with
exercise. She had always blamed her intolerance
to exercise on her sedentary lifestyle. However,
she recently joined a gym and after a week of
aerobics classes went to her physician, who
ordered a series of blood tests. The blood work
came back with abnormal results, leading to her
muscle biopsy and referral to the metabolic clinic.
22
Marie’s Blood Test Results
Levels (+ = normal, - = decreased levels, ++ = increased levels)
Blood Metabolic Panel
Blood Glucose
+
Glucose-6-phosphate
++
Fructose-6-phosphate
++
Fructose-1,6,bisphosphate
-
Glyceraldehyde-3-phosphate
-
1,3-bisphosphoglycerate
-
Pyruvate
-
ATP
-
Red Blood Cell Concentration
23
CQ#5: The muscle biopsy showed an
excess of glycogen in the muscle tissues.
What is the most likely enzyme deficiency?
A. Pyruvate kinase
B. Hexokinase
C. Phosphofructokinase
D. Triosephosphate Isomerase
E. Aldolase
24
CQ#6: If you wanted to test red blood cells
for their ability to complete glycolysis, what
compound would you try to detect?
A. Fructose-6-phosphate
B. Aldolase
C. Pyruvate
D. Phosphofructokinase
E. Dihydroxyacetone phosphate
25
CQ#7: If you had some of Ann’s red blood
cells in a test tube, what compound could
you add to enable these cells to produce
pyruvate?
A. Glucose
B. Fructose-1,6,bisphosphate
C. Insulin
D. Glucagon
E. Fructose-6-Phosphate
26
Patient #3: Leo is a 25-year-old accountant. He is
not anemic, has perfect teeth, and has experienced
two fainting spells recently. One happened after
accidentally drinking a non-diet soda; the other after
eating “naturally sweetened” fruit salad. His dentist
referred him to Dr. Saccharo’s clinic. Dr. Saccharo
suspects Leo may have hereditary fructose
intolerance (HFI) and has recommended a closely
monitored fructose tolerance test. Leo is scheduled
for the test and an observation bed at the local
hospital that afternoon.
27
Hereditary Fructose Intolerance
• HFI is a recessive disorder where individuals with HFI lack a
functional copy of Aldolase B.
• Individuals with HFI normally become symptomatic with
weaning – as babies switch from breast-milk with lactose to
formula with sucrose or sorbitol. The HFI may go
undetected until adulthood, with adults self-regulating their
diet to be low in sugars.
• They are able to metabolize glucose, because they have
Aldolase A and C. However, in the liver, it is Aldolase B that
breaks down fructose-1-phosphate, sending
dihydroxyacetone phosphate and glyceraldehyde-3
phosphate to glycolytic pathway. Since HFI patients lack
Aldolase B, they are unable to break down fructose.
28
Fructose metabolism
– In the liver
Inside the
cell
Fructose 1Phosphate
Inhibits
Glycogen
phsphorylase
Blood stream
Glycogen
Fructokinase
Fructose
Aldolase B
DHAP
G3P
Glucose-6phosphate
Dihydroxyacetone
phosphate
(DHAP)
Glyceraldehyde-3phosphate
(G3P)
29
CQ#8: Glycogen phosphorylase is an
enzyme required to release the glucose
stored as glycogen. A person with HFI
who ingested fructose would have:
A.
B.
C.
D.
E.
high levels of blood fructose.
high levels of blood fructose-1-phosphate.
high levels of blood glucose.
Low levels of blood fructose.
low levels of blood glucose.
30
Regulation of Glycolysis
• Glycolysis is reversible. Most of the enzymes can work
to break down glucose (gycolysis) or build more glucose
(gluconeogenesis).
• Products of a particular enzymatic reaction will generally
inhibit that enzyme.
• Several points of control exist. For example:
– Hexokinase is inhibited by glucose-6-phosphate and
activity is boosted by free phosphate.
– Phosphfructokinase is inhibited by ATP and Fructose1,6,-bisphosphate
– Pyruvate kinase is inhibited by ATP, but activated by
Fructose-1,6,bisphosphate and phosphoenolpyruvate.
31
The Afternoon at the Hospital
When you arrive at the nurse’s station to
start your shift the next day, you discover
that Leo just started his fructose tolerance
test.
You see Dr. Saccharo behind the desk,
where he’s overseeing the test.
32
Fructose Tolerance Test Blood
Glucose Levels
100
Glucose (mg/dL serum)
90
80
70
60
50
40
0
15
30
45
60
Time After Fructose (250 mg/kg) Injection
(Minutes)
33
CQ#9: What was the effect of giving Leo
intravenous fructose?
A. A rise in blood glucose levels.
B. A decrease in blood glucose levels.
C. An increase in glycogen stores.
D. A rapid drop in blood pressure.
34
Explanation of Test Results
• The lack of Aldolase B causes a build up
of fructose-1-phosphate.
– This will trap phosphate in the liver.
– Fructose-1-phosphate will inhibit glycogen
phosphorylase.
• Glycogen phosphorylase is required for
the liver to break glycogen into glucose-6phosphate.
35
Complications
• As soon as the test is completed, Leo orders
and drinks a cup of tea. However, after half a
cup, Leo passes out.
• Dr. Saccharo orders two blood sugar checks.
Leo’s blood glucose was 50 mg/dL; 15 minutes
later, it was down to 40 mg/dL.
• With the last test result, Dr. Saccharo rushes
into Leo’s room and injects something into
Leo’s IV. Within minutes, Leo is back to
normal.
36
CQ#10: What did Dr. Saccharo inject into
the intravenous line to rescue Leo?
A. Insulin
B. Glucagon
C. Glucose
D. Fructose
E. Phosphofructokinase
37
Leo’s Prognosis
The dramatic results of the fructose
tolerance test confirmed that Leo had
hereditary fructose intolerance. Dr.
Saccharo has put Leo on a restricted diet
with a plan for regular follow-up visits.
38
You leave your first day of the
internship thinking, I wonder
what’s on the agenda
tomorrow?
39