Glycation in Demetalated Superoxide Dismutase 1 Prevents Amyloid Aggregation and Produces Cytotoxic Ages Adducts

Abstract

Superoxide dismutase 1 (SOD1) has been implicated with familial amyotrophic lateral sclerosis (fALS) through accumulation of protein amyloid aggregates in motor neurons of patients. Amyloid aggregates and protein inclusions are a common pathological feature of many neurological disorders in which protein aggregation seems to be directly related to neurotoxicity. Although, extensive studies performed on the aggregation process of several amyloidogenic proteins in vitro allowed the identification of many physiological factors involved, the molecular mechanisms underlying the formation of amyloid aggregates in vivo and in pathological conditions are still poorly understood. Post-translational modifications are known to affect protein structure and function and, recently, much attention has been devoted to the role played by non-enzymatic glycation in stimulating amyloid aggregation and cellular toxicity. In particular, glycation seems to have a determining role both in sporadic and familial forms of ALS and SOD1 has been shown to be glycated in vivo The aim of this study was to investigate the role of glycation on the amyloid aggregation process of both wild-type SOD1 and its ALS-related mutant G93A. To this aim, the glycation kinetics of both native and demetalated SOD have been followed using two different glycating agents, i.e., D-ribose and methylglyoxal. The effect of glycation on the structure and the amyloid aggregation propensity of native and ApoSOD has been also investigated using a combination of biophysical and biochemical techniques. In addition, the effect of SOD glycated species on cellular toxicity and reactive oxygen species (ROS) production has been evaluated in different cellular models. The results provided by this study contribute to clarify the role of glycation in amyloid aggregation and suggest a direct implication of glycation in the pathology of fALS.

Analyzing structural alterations of mitochondrial intermembrane space superoxide scavengers cytochrome-c and SOD1 after methylglyoxal treatment

“…In our study, MGO concentrations of 100 μM and 1 mM are effectively glycating SOD1. At the latter MGO concentration, SOD1 oligomerization is observed, confirming an earlier result by the Iannuzzi lab [24] (Fig 6).…”

“…In contrast, glycation with a high level of MGO (1 mM) leads to the appearance of an additional maximum at approximately 66˚C and a slight shift of the main maximum to 90˚C. The former peak might be based on a de-metallated form of the protein (i. e., apoSOD1) that is present in the sample whereas the latter indicates a slightly stabilizing effect of MGO on SOD1 which was reported previously [24] and which we could demonstrate by immunodetection ( Fig 2B). The pronounced aggregation of SOD1 Table 2.…”

“…In vitro glycation experiments using mostly MGO and a target molecule have been instrumental in elucidating the chemistry of the reactions that dicarbonyls can undergo [21][22][23][24][25][26][27]. In this study, we focused on two proteins which play an important role in the detoxification of the reactive oxygen species (ROS) superoxide ( .…”

Vanillin Affects Amyloid Aggregation and Non-Enzymatic Glycation in Human Insulin

“…Cells in culture medium without protein and in the presence of curcumin and vanillin at the tested concentrations served as control. In the AGE experiments, before cells exposure, insulin glycated in the presence of 0.5 M D-ribose for 8 days was subjected to dialysis in sterile conditions to remove the free glycating agent 89 .…”

The Role of Metal Binding in the Amyotrophic Lateral Sclerosis-Related Aggregation of Copper-Zinc Superoxide Dismutase

“…Indeed, SOD1 is an extremely stable protein when fully assembled, but it is rather unstable and prone to aggregation in its metal-free form. Many studies have shown that wild-type human SOD1, when lacking both its metal ions, forms large and stable amyloid-like protein aggregates under physiological conditions of pH and temperature thus suggesting that metal binding could play a key role in the in vivo aggregation process of SOD1 [ 15 , 24 , 25 , 26 , 27 , 28 , 29 ]. Moreover, it has been reported that also some SOD1 mutants, many of them related to familial ALS, form amyloid aggregates both in vitro and in vivo and that demetalation is the key factor for aggregation [ 15 , 25 , 29 , 30 , 31 , 32 , 33 , 34 , 35 , 36 ].…”