Biological Polyanions Promote Aggregation Of Wild-Type And Mutant SOD1

Amyotrophic lateral sclerosis (ALS) is a common fatal neurodegenerative diseases. Increasing evidence show that aggregation of wild-type Cu/Zn superoxide dismutase (SOD1) and its mutations is one of the most important causes in ALS, and changing in structures and properties from oxidative damage, mutation or other reasons would result in SOD1aggregation. It is now generally believed that soluble aggregates or oligomers are more toxic than mature aggregates. DNA is one of the most abundant polyanion in vivo. We have found that DNA could accelerate wtSOD1aggregation in acid condition and oxidative damage SOD1in neutral condition. Heparin is a highly sulfated linear polyanion species and associate with virtually all amyloid deposits in vivo. Research in heparin and DNA as a representative of polyanion promoting SOD1and its mutants aggregation may provide some experimental basis in understanding the pathology of ALS.We have investigated the interaction between polyanion and SOD1or A4V through fluorescence spectra, RALS, DLS, CD, TEM, gel electrophoresis and cell toxic experiment, main results are as follows:Firstly, polyanion could accelerate the aggregation of SOD1or A4V dependent on the pH value, and wtSOD1needs a lower pH to achieve the same level, indicating that A4V has a higher tendency to aggregate. Histidine plays an important role in polyanion binding SOD1, which may be the main reason for the pH dependence. These results indicate that electrostatic interaction may be dominant in the interaction.Secondly, By using the tryptophan fluorescence quenching and fitting method, we have acquired the quenching and binding constants of polyanion to SOD1and found that the interaction between polyanion and SOD1showing the laws of LMWHep≥pBR322DNA～ctDNA>λDNA. As similar concentration, LMWHep would inhibit the combination of DNA to SOD1in pH≤4.0and the inhibition will reduce with increasing pH values.Thirdly, the aggregate morphology of A4V interaction with heparin or DNA is obviously different. A4V-heparin simple is loose, but A4V-DNA is more compact and rich in fibrous structure and different types of DNA would also lead to different morphological differences. Both heparin and DNA could gradually reduce the peaks at 208nm of SOD1CD spectrum, but as the pH value increased, DNA will induce a higher change in the secondary structure of wtSOD1than LMWHep, implying there is weak non-electrostaticeffect interaction between DNA and SOD1.At last, the supernatant of A4V-LMWHep or pBR322DNA reaction system show higher cytotoxic than mature aggregates due to the presence of soluble aggregates or oligomers.