This document summarizes a presentation on interactions between GBE1 and membranes and their implications for adult polyglucosan body disease (APBD). It discusses how GBE1 binding to membranes may be altered by mutations and age-related lipid changes, linking lipids to APBD pathology. Studies showed GBE1 localizes to cell membranes and binds model membranes. Supplementation with triheptanoin or hydroxylated triglyceride drugs lowered the membrane phase transition temperature, suggesting these lipids and drugs could be used to treat APBD by modulating membrane structure. Future research plans include further investigating wild type and mutant GBE1 interactions with membranes and the effects of triglyceride molecules on these interactions.

1. 10th Meeting of the APBDRF. New York City, 2013
GBE1 interactions with membranes:
Implications in APBD
Rafael Álvarez, David J. López, Maitane Ibarguren and Pablo V. Escribá
University of the Balearic Islands, Palma de Mallorca, Spain.

2. Glycogen storage disease types
Corpora amylacea in the brain cortex
of a 56-yr man. A) PAS-hematoxylin
stain; bar = 20 µm. B) Hematoxylineosin stain; bar = 40 µm.

3. Considerations about etiology and therapy
1. The late onset of APBD suggests a critical cellular event in the
etiology of APBD different from mutations.
2. Decline of DHA and PE with age suggests that GBE1-lipid
interactions could be associated with APBD.
3. Nutraceutical supplementation with Triheptanoin links lipids
with APBD and suggests that MLT approaches could be used to
treat this condition.
4. Some authors suggest that misfolded GBE1 may cause APBD. We
believe that misfolding could alter the cell localization of GBE1.

4. GBE1 (OriGene) binding
to model membranes

5. GBE1 in U118 cells
Confocal microscopy of GBE1 in U118 cells. Cells were stained with Hoechst 33342 (blue, nucleus) and
with an anti-GBE1 antibody (red). Bar=10 μm.

6. GBE1 amino acid sequence
WT Glucan (1,4-alpha-) branching enzyme 1 ML Databases
WT Glucan (1,4-alpha-) branching enzyme 1 OriGene Tech. Co.

7. GBE1 amino acid sequence
WT Glucan (1,4-alpha-) branching enzyme 1 Escribá
Y329S Glucan (1,4-alpha-) branching enzyme 1 Escribá

8. GBE1 IN Sf9 MEMBRANES

9. GBE1 in Sf9 cell Cytosol and purification

10. GBE1 bionding to model membranes

11. APBD: Drug design and screening
Tonset (ºC)
Tm (ºC)
ΔH
(Kcal/mol)
Tonset (ºC)
TH (ºC)
POPE
22.8 ± 0.2
23.6 ± 0.2
4.42 ± 0.03
68.3 ± 0.4
69.1 ± 0.5
0.42 ± 0.03
-----
+ 1 mol% Triheptanoin
21.4 ± 0.4
23.1 ± 0.2
5.90 ± 0.22
61.0 ± 0.9
62.5 ± 0.7
0.50 ± 0.06
↓7 ºC
+ 1 mol% TGM7
21.4 ± 0.1
22.7 ± 0.2
5.88 ± 0.97
66.6 ± 0.6
68.3 ± 0.6
0.51 ± 0.04
↓1 ºC
+ 1 mol% TG140
22.3± 0.5
24.4 ± 0.3
4.41 ± 0.05
58.7 ± 1.7
63.8 ± 0.7
0.23 ± 0.06
↓5 ºC
+ 1 mol% TG141
22.3 ± 0.5
23.5 ± 0.2
5.12 ± 0.12
61.0 ± 1.4
64.7 ± 0.8
0.21 ± 0.10
↓4 ºC
+ 1 mol% TG160
22.7 ± 0.3
23.8 ± 0.2
10.2 ± 0.19
58.2 ± 0.2
58.6 ± 0.2
1.21 ± 0.24
↓10 ºC
+ 1 mol% TG161
22.6 ± 0.3
23.5 ± 0.2
4.2 ± 0.10
61.3 ± 1.1
64.5 ± 0.7
0.15 ± 0.05
↓5 ºC
+ 1 mol% TG180
22.8 ± 0.3
23.8 ± 0.2
5.52 ± 0.19
62.7 ± 0.2
63.6 ± 0.4
0.84 ± 0.05
↓5 ºC
+ 1 mol% TG181
22.6 ± 0.3
23.3 ± 0.2
4.55 ± 0.09
58.5 ± 4.0
60.8 ± 3.3
0.24 ± 0.11
↓8 ºC
+ 1 mol% TGM1
22.7 ± 0.2
23.8 ± 0.3
5.78 ± 0.08
64.0 ± 0.6
66.0 ± 0.9
0.55 ± 0.06
↓3 ºC
+ 1 mol% TG182
22.2 ± 0.2
23.4 ± 0.2
5.52 ± 0.07
52.7 ± 5.2
61.6 ± 1.2
0.36 ± 0.11
↓8 ºC
+ 1 mol% TGM2
21.5 ± 0.2
23.0 ± 0.2
5.78 ± 0.07
56.0 ± 2.1
62.0 ± 1.1
0.67 ± 0.24
↓7 ºC
+ 1 mol% TG204
22.2 ± 0.4
23.4 ± 0.2
5.68 ± 0.22
58.9 ± 0.2
63.2 ± 0.4
0.42 ± 0.03
↓5 ºC
+ 1 mol% TGM4
21.9 ± 0. 2 23.2 ± 0.2
3.55 ± 0.22
60.4 ± 1.5
64.6 ± 0.8
0.37 ± 0.09
↓5 ºC
+ 1 mol% TG226
22.5 ± 0.2
5.73 ± 0.10
59.5 ± 2.3
63.7 ± 1.9
0.40 ± 0.07
↓5 ºC
23.5 ± 0.2
ΔH (Kcal/mol) Effect on TH
Table 1.mol% TGM6 transition temperatures of POPE mixed with 59.5 ± 1.3
+ 1 Enthalpies and
22.0 ± 0.1 23.3 ± 0.2 5.54 ± 0.12 1 mol% hydroxylated and non-hydroxylated triacylglycerides
64.9 ± 0.9
0.48 ± 0.13
↓4 ºC

12. Membrane structure modulation

13. POPE
15ºC
HΙΙ
80ºC
Lβ
Lα
18ºC

14. POPE

15. POPE
+ 5 mol% TG 2OHOA
Lamellar-to-hexagonal transition temperature (T H)
decreases drastically
15ºC
80ºC
18ºC

16. POPE + 5 mol% TG 2OHOA
Lamellar-to-hexagonal transition temperature (T H)
decreases drastically

17. APBD FUTURE RESEARCH
1.
2.
3.
4.
5.
6.
Determination of our GBE1 proteins (WT and Y329S) activity.
Protein production and purification.
Investigate of WT and Y329S GBM1 with different types of membranes
Investigate the role of TGMs and other LP drugs on these interactions.
Get a proof of principle for one oe more molecules.
(GBE1 structure).