1. Hereditary fructose intolerance and the aldolase B gene
Maddy Ford
May 8th, 2014

2. What is HFI?
Inability to process fructose

3. Personal perspective on HFI
“It’s such a part of my life, I don’t think about it much anymore” –Becky Heiser, diagnosed with HFI

4. What causes HFI? Defective aldolase B enzyme A174D A149P L256P R59OP
N334K
A337V
Glycolytic domain
Low complexity region (LCR)
Molecular Function
Biological Process
Cellular Component

5. How does aldolase B protein differ between homologs?
LCR
100%
99%
96%
76%
65%
58%
Human
Chimpanzee
Mouse
Zebrafish
Fruit fly
Nematode
Glycolytic domain
364
Glycolytic domain
LCR
364
Glycolytic domain
301
Glycolytic domain
301
LCR
Glycolytic domain
LCR
361
Glycolytic domain
LCR
366

6. The protein phylogeny of aldolase B
Human
Chimpanzee
Mouse
Zebrafish
Fruit fly
Nematode

7. Knowledge gap: Are the C-terminus or LCR important for human aldolase B function?
100%
99%
96%
76%
65%
58%
Human
Chimpanzee
Mouse
Zebrafish
Fruit fly
Nematode
364
LCR
364
301
301
LCR
LCR
361
LCR
366
Hypothesis: The C-terminus is important because it contains common mutation sites, the LCR is important for enzymatic folding.

8. Specific Aim 1: To determine the importance of the C-terminus in fructose metabolism
Why: Understanding aldolase B structure and function
How: RNAi
LCR
Hypothesis: knockout flies would be unable to process fructose, eventually die

9. Aim 1 expected results
-/-
LCR WT

10. Is the LCR important for fructose metabolism?
100%, 364 a.a.
99%, 364 a.a.
96%, 301 a.a.
76%, 301 a.a.
65%, 361 a.a.
58%, 366 a.a.
Human
Chimpanzee
Mouse
Zebrafish
Fruit fly
Nematode
LCR
LCR
LCR
LCR
LCR
LCR: ASALAAWGGKAANKKA

11. Hypothesis: all LCR knockouts will be unable to process fructose
Specific Aim 2: To determine the LCR’s importance in fructose metabolism
Why: Understanding aldolase B structure and function
How: RNAi
LCR
LCR
LCR
LCR
LCR
LCR
LCR
LCR
Hypothesis: all LCR knockouts will be unable to process fructose

12. LCR could be important for enzymatic folding
Aim 2 expected results
LCR
LCR
LCR
LCR
LCR
LCR
LCR
LCR
LCR could be important for enzymatic folding

13. Can we find a substitute for aldolase B?
Aldolase B is unstable  Find a substitute to mimic its action

14. Specific Aim 3: To identify chemicals that mimic aldolase B, and what genes they affect.
Why: Investigating HFI drug possibilities
How: Chemical genetics
Hypothesis: Drugs similar to aldolase B in structure and function will rescue WT phenotype, increase glycolytic gene expression

15. Aim 3 expected results Glycolytic gene expression Effective drug
Ineffective drug
Glycolytic gene expression

16. Conclusions
Further testing of the aldolase B gene and protein could provide new insight on enzyme activity Chemical genetics could yield a drug to allow individuals with HFI to consume fructose without experiencing symptoms
LCR

17. Future directions
Determine the importance of Alanine within the LCR: ASALAAWGGKAANKKA Researching similarity between mouse and zebrafish Further chemical testing

18. References
1. Cox, T.M. (1994). Aldolase B and fructose intolerance. The FASEB Journal, 8(1), James, C.L., Rellos, P., Ali, M., Heeley, A.F. & Cox, T.M. (1996). Neonatal screening for hereditary fructose intolerance: frequency of the most common mutant aldolase B allele (A149P) in the British population. Journal of Medical Genetics, 33(10), 837– Kajihara, S., Mukait, T., Arai, Y., Owada, M., Kitagawa, T. & Hori, K. (1990). Hereditary Fructose Intolerance Caused by a Nonsense Mutation of the Aldolase 4. Understand Dairy Sensitivity: Overview. Lactaid. Accessed 13 April < 5. Ekker, S.C., and Larson, J.D. (2001). Morphant technology in model development systems. Genesis 30:89– Miller, D, Hannon, C, and Ganetzky. (2013). A mutation in Drosophila aldolase causes temperature-sensitive paralysis, shortened lifespan, and neurodegeneration. Journal of Neurogenetics. 26 (3-4): doi: / Thurston, J.H., Jones, E.M., Hauhart, R.E. (1974). Decrease and Inhibition of Liver Glycogen Phosphorylase After Fructose. Diabetes, 23 (7), doi: /diab NCBI BLAST. Homo sapiens chromosome 9, GRCh37.p13 Primary Assembly compared to Mus musculus aldolase B, fructose-bisphosphate, mRNA (cDNA clone MGC:36362 IMAGE: ), complete cds.

19. Image references
Baby with fruit.
Human liver.
Fructose.
Glucose.
Cytosol.
Aldolase B in glycolysis.
Clustal Omega logo.
Clustal Omega sequence aligntment.
Drosophila.
Drosophila on fruit.
Strawberry.
C. elegans.
Fruit fly on fruit.
Lactaid.
Morpholinos.
Fly screen.
Business cat.

20. Questions? Thanks for listening.