Monday at the Metabolic Clinic

Adventures in Glycolysis

by

Nancy Boury

Iowa State University

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BackgroundYou 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.

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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.

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Please get into your groups of four and take out your clickers

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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)

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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

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Dr. Saccharo’sReview 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.

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Dr. Saccharo’sReview 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 potentially die in a matter of hours.

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Fasting- Glucose

Levels Low

Fasting- Glucose

Levels Low

Pancreas Releases Glucagon

Pancreas Releases Glucagon

Liver Glucagon

Receptors

Liver Glucagon

Receptors

Liver Releases

Glucose into the Blood

Liver Releases

Glucose into the Blood

Fed – Blood Glucose

Levels High

Fed – Blood Glucose

Levels High

Pancreas Releases

Insulin

Pancreas Releases

Insulin

Body Cells have Insulin Receptors

Body Cells have Insulin Receptors

Body Cells Take Up

Glucose from the Blood

Body Cells Take Up

Glucose from the Blood

Glucose Levels

Normal

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Dr. Saccharo’sReview 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.

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• 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.

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Please trade papers within your groups and check each other’s homework

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Embden-Myerhof Pathway

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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?

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“If glycolysis is needed to use glucose, how can a person live without all the

enzymes needed for glycolysis?”

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Jamie, one of your fellow interns, is bothered because he knew the clinic frequently treats people with enzyme deficiency disorders. He asks Dr. Saccharo:

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.

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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

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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.

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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.

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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.

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 -

Levels (+ = normal, - = decreased levels, ++ = increased levels)

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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

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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.

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Marie’s Blood Test ResultsLevels (+ = normal, - = decreased levels, ++ = increased levels)

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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 -

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

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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

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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

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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.

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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.

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Fructose metabolism – In the liver

Fructokinase

Glyceraldehyde-3-phosphate

(G3P)

Dihydroxyacetone phosphate

(DHAP)

Fructose

Fructose -1-

Phosphate

Blood streamInside the cell

Glycogen

Glucose-6-phosphate

Aldolase B

Inhibits

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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. high levels of blood fructose.

B. high levels of blood fructose-1-phosphate.

C. high levels of blood glucose.

D. Low levels of blood fructose.

E. low levels of blood glucose.

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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 Fructose-

1,6,-bisphosphate– Pyruvate kinase is inhibited by ATP, but activated by

Fructose-1,6,bisphosphate and phosphoenolpyruvate.

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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.

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Fructose Tolerance Test Blood Glucose Levels

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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.

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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-6-phosphate.

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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.

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CQ#10: What did Dr. Saccharo inject into the intravenous line to rescue Leo?

A. Insulin

B. Glucagon

C. Glucose

D. Fructose

E. Phosphofructokinase

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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.

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You leave your first day of the internship thinking, I wonder

what’s on the agenda tomorrow?

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