Kinetic modelling helps to understand oxidative stress caused by an antioxidant

Transcription

1 Kinetic modelling helps to understand oxidative stress caused by an antioxidant Axel Kowald Max Planck Institute for Molecular Genetics, Berlin

2 Trisomy1 (Down Syndrome) Three copies of chromosome 1 First described 1866 by J.. Down Genetic basis discovered 1959 Prevalence ca. 1:800 Mental retardation and characteristic facial features Overexpression of the ca. 50 genes of chromosome 1 Genes with possible relevance for Down Syndrome: SOD, CO6A1, ETS, CAF1A, CBS, DYRK, CRYA1, GART, IFNAR

3 Superoxide Dismutase O H O H O i SOD cat / GPX do dt i = k k SOD O i 1 O i,ss = k k1 SOD dh O dt = k SOD O k cat H O i 3 HO,ss = k 3 k1 cat 3

4 Negative Effects Increased lipid peroxidation in Trisomy1 Increased lipid peroxidation in cell cultures Increased mortality in transgenic D. melanogaster ipid peroxidation, diminished prostaglandin synthesis and serotonin uptake in transgenic mice. Bacteria overexpressing SOD showed increased paraquat sensitivity. 4

5 Explanations The reaction of H O with CuZnSOD leads to hydroxyl radical generation. O *- radicals might reduce membrane damage by acting as chain breakers. 5

6 ipid Peroxidation Chain Reaction 6

7 Explanations The reaction of H O with CuZnSOD leads to hydroxyl radical generation. O *- radicals might reduce membrane damage by acting as chain breakers. SOD cycles between a reduced and oxidised state. At low O *- levels the intermediates might interact with other redox partners and increase the superoxide reductase activity of SOD. 7

8 Reactions SOO SOR 8

9 Standard Simulation HO Om CuSODCu1SOD H Om HO H CuSOD Cu1SOD 8k ,k ,k ,k4 10 5,k5 10 4, k6 1, k ,k9 10 6, k10 000, k , k , k13a , k13b , k ,k18 14, k , SOD 10 5, cat 10 5 < 9

10 Increasing SOR Activity Cu1SODCuSOD H HO Om µ10-6 4µ µ µ SOD Om HO H CuSOD Cu1SOD k3b = 10*k3a 10

11 Increasing SOO Activity Cu1SODCuSOD H HO Om µ10-6 4µ µ µ SOD Om HO H CuSOD Cu1SOD 10*k3b = k3a 11

12 SOO/SOR Results In principle the SOO/SOR idea works. However, it is not clear why in experiments SOR activity is always greater than SOO activity. Alternative redox partners have only been demonstrated for CuZnSOD, not MnSOD. 1

13 Membrane Damage Termination 5 µ 10-6 CuSOD Om 4 µ µ 10-6 µ µ 10-6 Cu1SOD H µ µ µ µ SOD Om HO H CuSOD Cu1SOD * remains constant throughout the entire range * and H increase for small SOD values, but are constant otherwise. Why are the concentrations of * and * not increasing with increasing SOD? 13

14 Simplified Modell k 4 * O i,ss = k k1 + SOD k 1 k O *- k 3 * k 3b i ss = k k 4 SOD i ss k k3b k1 k = + k k k (k + SOD) * doesn t change because it is completely independent of SOD and O *-. If the concentration of O *- changes, membrane damage initiation changes by the same factor as the termination (for a given k 4 ). Without the back reaction k 3b, levels of * should decrease continously with increasing SOD, and this can be confirmed by disabling this reaction in the full modell. => Chain breaker idea does not work!! 14

15 Alternative Pathway Mechanism CuSOD Cu1SOD H HO Om µ µ µ µ SOD Om HO H CuSOD Cu1SOD Simulation with full modell and SOO equal to SOR (no effect) 15

16 Simplified Modell O H O H O k1 i k SOD k3 cat k 10 other reactions O HO i,ss,ss = = k1 k + k SOD 10 k1 k SOD k + k SOD k cat ( ) 10 3 If there are alternative reaction pathways for the superoxide radical, increasing SOD channels more of the available O *- into the H O generating branch and can thus increase oxidative stress. This principle is independent of the SOD type (CuZnSOD, MnSOD). k 10 does not only represent membrane damage, but A other reactions of O *-. Therefore k 10 can be much larger than the value used in the modell and so the resulting effect on H O can also be much larger. 16

17 Summary Overexpression of the antioxidant SOD leads to increased oxidative stress. We created a mathematical model to test some possible explanations. Only one proposed idea (SOD cycling) works as suggested under certain conditions. Studying the model we discovered an additional, more general, mechanism that might account for the increased oxidative stress. 17