Case 5

A 17 month old girl was admitted to the pediatric clinic due to prolonged anemia.

Aside from being pale, there were no other complaints.

Parents: both healthy apart from a gall bladder removal procedure that the father
underwent at age 28.

The girl’s hemoglobin values were always at the lower range of normal. Immediately upon
birth, she was jaundiced and anemic and received two blood transfusions. Since then,
despite iron therapy, her hemoglobin values were under 9 g/l and her reticulocyte counts 4-
11% (normal up to 1%).Bilirubin levels were high. In spite of the apparent anemia, the girl
felt good and was as active as her age-matched friends.

Blood was drawn for the following tests, all performed on whole blood lysate:

• PFK-1 activity in the absence and presence of F26BP:x1.8 higher than control
• All glycolysis metabolites were high, including 2,3-DPG and especially PEP (x3.5 from
control).
• Pyruvate kinase activity was measured as a function of ADP concentration in the
absence and presence of FDP

Questions

1. What is the diagnosis of this patient?

2. Explain what is the significance of glycolysis and PPP pathways in RBCs? How is it
different from epithelial or adipocyte cells?
3. Which enzyme is defected?
4. Why do we see an increase in PFK1 activity in the patient?
5. What is the physiological significance of 2,3-DPG? How does it relate to the patient's
symptoms?
Case 6

A fasting blood test was taken from 6 years old boy who is generally healthy. Blood glucose
was found to be 7.8 mmol/l and HbA1C 6.8%. Based on his family history genetic analysis
was performed and he was found to carry heterozygous inactivating glucokinase mutations.
This mutation is known to cause the autosomal dominantly inherited MODY2 subtype of
maturity-onset diabetes of the young.

It is known that another mutation in glucokinase can cause persistent hyperinsulinemic

hypoglycemia in infancy

Question:

Explain the two situations

Case 7

In 2013 a paper was published in BBRC ( Biochem Biophys Res Commun. 2013 Feb
8;431(2):367. ) Altered allosteric regulation of muscle 6-phosphofructokinase causes Tarui
disease. The abstract of this paper is enclosed:

Abstract

Tarui disease is a glycogen storage disease (GSD VII) and characterized by exercise
intolerance with muscle weakness and cramping, mild myopathy, myoglobinuria and
compensated hemolysis. It is caused by mutations in the muscle 6-phosphofructokinase
(Pfk). Pfk is an oligomeric, allosteric enzyme which catalyzes one of the rate-limiting steps of
the glycolysis: the phosphorylation of fructose 6-phosphate at position 1. Pfk activity is
modulated by a number of regulators including adenine nucleotides. Recent crystal
structures from eukaryotic Pfk displayed several allosteric adenine nucleotide binding sites.
Functional studies revealed a reciprocal linkage between the activating and inhibitory
allosteric binding sites. Herein, we showed that Asp(543)Ala, a naturally occurring disease-
causing mutation in the activating binding site, causes an increased efficacy of ATP at the
inhibitory allosteric binding site. The reciprocal linkage between the activating and
inhibitory binding sites leads to reduced enzyme activity and therefore to the clinical
phenotype. Pharmacological blockage of the inhibitory allosteric binding site or highly
efficient ligands for the activating allosteric binding site may be of therapeutic relevance for
patients with Tarui disease.

Question:

As a consultant for drug company, suggest ligands to treat these diseases in the future.