Transcript Fumarase Deficiency

PHM142 Fall 2016
Instructor: Dr. Jeffrey Henderson
Fumarase Deficiency
Dharen Ratnakumar, Arvind
Grewal, Amir Khela, Rikesh Patel
September 27, 2016
What is Fumarase Deficiency?
• Autosomal recessive metabolic disorder
• Extremely Rare Disorder
• Approximately 100 cases reported
https://en.wikipedia.org/wiki/Fumarase_deficiency#/media/File:Autorecessive.svg
What is Fumarase Deficiency?
Deficiency of the enzyme fumarate hydratase (fumarase)
Found in Kreb cycle
https://en.wikipedia.org/wiki/Fumarase#/media/File:Reaction1.png
Fumarase Mechanism of Action (Hydration)
https://en.wikipedia.org/wiki/Fumarase#/media/File:FumaraseMechanismE1CB.png
Cellular
Respiration
http://florianbrnada.weebly.com/uploads/1/7/9/4/17946175/5696532.jpg?597
Kreb’s Cycle
https://upload.wikimedia.org/wikipedia/en/4/45/Citric_acid_cycle_noi.JPG
Fumarase
-
Two Isoforms
- Cytosolic Form
- Mitochondrial Form
- Extended N-Terminus
- Identical subunits
-
Tetrameric Enzyme (54kDa each)
- Binding Site A (E315)
- Affinity for Fumarate, D-malate,
and Oxaloacetate
- Competitive Inhibitors: Glycine
- Binding Site B
- Lower affinity for same
substrates
http://www.ncbi.nlm.nih.gov/Structure/icn3d/full.html?showseq=1&mmdbid=66080&buidx=0
Fumarase Inhibitors
Competitive Inhibitors:
-
Maleic Acid: found in drugs like Carfenazine & Chlorpheniramine
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Glutaric Acid: by-product of Tryptophan catabolism
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D-Tartaric Acids: added to oral drugs to improve taste, such as cough
syrups
Molecular Genetics
FH gene located on the long arm of chromosome 1
17 FH gene mutations known to cause fumarase deficiency
● Mainly missense mutations
Parents of affected child are unaffected obligate heterozygotes
Clinical Characteristics
The clinical presentation of Fumarase Deficiency in a patient is characterized
by:
Encephalopathy
Major cognitive deficiencies
Epileptic seizures
Irregular facial features and brain malformation
Clinical Characteristics
Brain Imaging
● Most common finding is a small brain
○ Cerebral atrophy
○ Ventriculomegaly
Clinical Course
● Many individuals do not survive infancy
● Death due to secondary complications (eg. respiratory failure)
● Patients unable to feed
● A few cases have lived into early adulthood
Clinical Diagnosis
Increased concentration of Fumaric Acid in the urine.
Confirmation of diagnosis through identification of deficient fumarate
hydratase enzyme activity.
Fibroblasts
Lymphoblasts
White Blood Cells
Enzyme activity of affected individuals usually 10% of the control group.
Genetic testing of FH gene
Sequence analysis to determine mutant alleles.
Most frequent mutant allele is c.1431_1433dupAAA
Amplification of sequence through qPCR.
Genetic Testing
Sample of DNA taken.
Denaturation of DNA followed by the primer annealing and extension.
Sequence comparison to the normal allele.
Sequence analysis of entire coding region
Deletion/Duplication analysis
Testing for Fumarase Deficiency often included in tests for other genetic
disorders.
E.g. Comprehensive Epilepsy NGS Panel, includes Fumarase Deficiency as
well as 3052 other conditions.
Treatment
Extent of Deficiency
MRI and evaluation by neurologist.
Management of epileptic seizures
Numerous medications but not the ketogenic diet.
Nutritional treatment
Management of lethargy and other symptoms associated with the encephalopathy.
Physical therapy
Management of the physical defects that can lead to such diseases as Scoliosis.
Summary
•
A rare autosomal recessive metabolic disorder leading to the deficiency of the enzyme fumarase
•
Mitochondrial Fumarase converts Fumarate to Malate in Kreb’s cycle through hydration reaction
•
It is enzyme with four identical subunits (50kDa each, tetramer)
•
Binding site A with higher affinity than binding site B for Fumarate
•
17 known mutations to FH gene on chromosome 1 cause fumarase deficiency
•
Diagnosis through Fumarate Hydratase and FH Gene testing
•
No direct treatment for Fumarase deficiency, rather addressing the many symptoms
Summary
cont.
https://upload.wikimedia.org/wikipedia/en/4/45/Citric_acid_cycle_noi.JPG
References
Gellera, C.; Uziel, G.; Rimoldi, M.; Zeviani, M.; Laverda, A.; Carrara, F.; DiDonato, S. (March 1990). "Fumarase deficiency is an
autosomal recessive encephalopathy affecting both the mitochondrial and the cytosolic enzymes". Neurology. 40 (3 Part 1): 495–
499.
Coughlin, E. M., Christensen, E., Kunz, P. L., Krishnamoorthy, K. S., Walker, V., Dennis, N. R., ... & Ramesh, V. (1998). Molecular
analysis and prenatal diagnosis of human fumarase deficiency. Molecular genetics and metabolism,63(4), 254-262.
Pereira de Padua, R. A., & Nonato, M. C. (2014). Cloning, expression, purification, crystallization and preliminary X-ray diffraction
analysis of recombinant human fumarase. Acta Crystallographica Section F: Structural Biology Communications, 70(1), 120-122.
Bulku, A., Aleksanyan, N., & Berkmen, M. (2016). Oligomerization of wild-type fumarase and a mutant associated with fumarate
hydratase deficiency.The FASEB Journal, 30(1 Supplement), 665-6.
Massey, V. (1953). Studies on fumarase. 4. The effects of inhibitors on fumarase activity. Biochemical Journal, 55(1), 172.
Bartlett, J. M. S.; Stirling, D. (2003). "A Short History of the Polymerase Chain Reaction". PCR Protocols. Methods in Molecular
Biology. 226 (2nd ed.). pp. 3–6.