AN APPLE A DAY KEEPS

THE DOCTOR IN PAY

BCHM2972 Lecture 10 2006

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Presentation and history:• A 10-year old girl, Jane, has a long history of problems

after eating fruit or any foods containing sugar.• The symptoms include vomiting, stomach cramps,

trembling, sweating, dizziness, ~ convulsions. • 'sickly' since weaned from breast milk to infant formula

and solids. • OK foods? trial and error.

– glucose OK but she does not enjoy the taste.– cow's milk OK

physical development• Difficult to maintain sugar free diet.• teeth show no caries!

• Jane has presented at hospital tonight with violent convulsions which were preceded by stomach cramps and sweating.

• After soft drink and lollies at a friend's slumber party (peer pressure!)

• Refer to the handout of Test results

What is the likely cause of the sweats, convulsions, etc?

A Hypoglycemia

B Hyperglycemia

C Hypoinsulinemia

D Hyperfructosemia

Which ‘sugars’ is Jane reacting to?

A. glucose

B. sucrose

C. galactose

D. fructose

E. lactose

Which ‘sugars’ is Jane reacting to?

A. glucose

B. sucrose

C. galactose

D. fructose

E. lactose

• We know • foods with glucose OK• cows milk OK• FRUIT a problem• lollies and soft drinks too

monosaccharides

sucrose invertase fructose + glucose

in small intestineabsorbed to blood

so…a problem with fructoseinborn error of metabolism?

• no fructose in breast milk!

Aldolase B not expressed in infants

• long history• always ‘sickly since

weaning’• Why not since birth?

fructose tolerance test:infuse 200mg/kg fructose intravenously

measure blood levels of:–fructose–glucose–phosphorus–magnesium–uric acid

which do you expect to be abnormal?

confirmed blood test levels are in response to fructose

What the…?

• Jane has a problem eating fructose, but fructose clearance is normal !!

• Why is glucose ( symptoms) ?

• ……and why P and Mg?

• and uric acid and bilirubin??

• don’t these mean liver damage?

• It appears fructose is very efficiently taken up and trapped in tissues

• it's the metabolism of the fructose in the tissues that's the problem

So…..let’s review “sugar” metabolism

Breakage of the Phosphoanhydride Bond in ATP

nucleophile (donates e-)1st step

in g

lycolysis

Glucose

Fructose 6 P

GA 3 PDHAP

6

PP

gly

coly

sis

hexokinase

isomerase

ATP

phosphofructokinase

Aldolase A

P

pyruvate

CITRIC

ACID

CYCLE

electron transport

and OxPhos

ATP

~30 ATP

per glucose

Pyr deHydrATP

PHOSPHORYLATION

ISOMERISATION

PHOSPHORYLATION

‘CLEAVAGE’

Glucose

Fructose 6 P

GA 3 PDHAP

6

PP

gly

coly

sis

hexokinase

isomerase

ATP

phosphofructokinase

Aldolase A

P

pyruvate

CITRIC

ACID

CYCLE

electron transport

and OxPhos

ATP

Fructose

ATP

hexokinase affinity for fructose

ATP

in muscle, fat cells

ATP

Glucose

Fructose 6 P

GA 3 PDHAP

6

PP

Fructose

Fructose 1 P1

DHAPP

ATP

gly

coly

sis

fructokinase

ATP

Aldolase B

hexokinaseATP

pyruvate

CITRIC

ACID

CYCLE

electron transport

and OxPhos

ATP

Glyceraldehyde (GA)

in liverhexokinase has affinity for glucose but affinity for fructose. lots of glucose in

liver so instead of hexokinase, fructokinase adds P to trap fructose in

cells

What is the result of the first step in fructose metabolism?

A. phosphorylation of fructose

B. ATP used up to ph’late fructose

C. cellular energy is reduced

D. phosphate ‘trapped’ in fructose 1 P

E. all of the above

What is the result of a mutation in fructokinase?

A. can’t ph’late fructose fructose- 1P

B. reduced clearance of fructose from blood

C. blood fructose remains high

D. ATP not used to p’late fructose

E. all of the above

Can Jane’s liver trap fructose in the tissue (clearing it from blood)?

A. Yes

B. No2468

1012

-20

0 30 60 90 120time (min)

fru

cto

se (

mg

/dL

)

Does Jane have a mutation in fructokinase?

A. Yes

B. No

Her blood fructose does not rise rapidly after the fructose load because fructokinase is very good a mopping up and trapping fructose in the liver. And/or hexokinase in other tissues is very good at mopping up fructose into F6P

Which enzyme catalyses the second step of fructose catabolism?

A. hexokinase

B. fructokinase

C. phosphofructokinase

D. aldolase A

E. aldolase B

What is the result of a mutation in aldolase B?

A. fructose -1P builds up

B. ph’lation of F-1P uses up ATP

C. cellular energy is reduced

D. phosphate ‘trapped’ in fructose- 1 P

E. all of the above

ATP

Glucose

Fructose 1 P

1

ATP

gly

coly

sis

fructokinase

Aldolase B

ATP

pyruvate 30 ATP

Fructose

Why the Pblood?

A. P trapped in fructose 1-PB. P trapped in fructose 6-PC. P trapped in fructose 1, 6 -biPD. P trapped in glyceraldehyde 3PE. all of the above

How do we explain Jane’s results by a mutation in aldolase B?

Why the Pblood?

A. P trapped in fructose 1-PB. P trapped in fructose 6-PC. P trapped in fructose 1, 6 -biPD. P trapped in glyceraldehyde 3PE. all of the above

How do we explain Jane’s results by a mutation in aldolase B?

What are the consequences of Pblood?

• ADP + P ATP

low cellular energy (ATP)

cell damage /death

release of liver enzymes and bilirubin

Why the high Mg?

• ATP in complex with Mg2+

ATP Mg2+ released to blood

0

0.1

0.2

0.3

0.4

0.5

0 30 60 90 120

time (min)

mag

nes

ium

(m

g/d

L)

ATP, Mg2+, and +ve residues at an enzyme active site

Since all the negative charges in ATP are neutralized, ATP is readily approached by nucleophiles

Glucose

Fructose 6 P

GA 3 PDHAP

6

PP

gly

coly

sis

hexokinase

isomerase

ATP

phosphofructokinase

Aldolase A

P

pyruvate

CITRIC

ACID

CYCLE

electron transport

and OxPhos

ATP

ATP

ATP Fructose

AMP ADP

ADP

+

substrate level phosphorylation to compensate for ATP

PFK rate gylcolysis (important regulatory point)

glucose

NADH

glycolysis

NAD+

pyruvate

CITRIC

ACID

CYCLE

electron transport

and OxPhos

ATPAcCoA

PDHPFK

normally……

pyruvate

CITRIC

ACID

CYCLE

electron transport

and OxPhos

glucoseATP

NADH

glycolysis

AMP

PDHPFK

+

NAD+

AcCoA

In Jane’s liver, pyruvate is produced too fast to enter mitochondria for TCA (PDH is like the ‘plughole’)

NAD not regenerated

cell must regenerate NAD+ to keep glycolysis running

desperate attempt to make ATP by glycolysis (2 per glucose)

pyruvateglucoseglycolysis

lactate

NADH

NAD+

ATP

What processes normally lead to increased blood lactate?

A Decreased liver gluconeogenesis

B Increased liver glycolysis (This happens in Jane via stimulation of PFK by AMP)

C Increased muscle glycolysis

D Decreased PDH activity (would entry of substrates to Citric Acid Cycle)

E All of the above

all increase pyrlactate to regenerate NAD

Glucose

Fructose

Fructose 1 P

ATPfructokinase

Aldolase B

gly

cog

eno

lysi

s

glu

con

eog

enesis

glycogen phosphorylase

Fructose 1,6 BP

fructose 1,6 bisphosphatase

P

Why is glucose sooooo low? in addition to glycolysis….

What is [uric acid*] blood

indicative of?

A cellular 'energy charge'

B rate of deamination of adenosine

C in ATP

D in AMP

E All of the above

*not to be confused with urea! (product of amino acid catabolism)

AMP

P

inosine MP uric acid

P

H

H

H

uric acid is a product of purine base degradation

deamination

What problems can fructose cause for ‘normal’ people?

• does NOT stimulate insulin or leptin – regulators of energy intake and body adiposity

• bypasses phosphofructokinase – determines the rate of glycolysis (pyruvate)

• So….. fructose acetyl CoA

Lipogenesis liver TG

insulin sensitivity

VLDL

Hepatic fructose metabolism: A highly lipogenic pathway.

http://www.nutritionandmetabolism.com/content/figures/1743-7075-2-5-2-l.jpg

In Summary…

You should make a summary diagram yourselves! Show main control points, interconnections, and show

how the non-functioning enzymelab results