Noncovalent Interactions between Superoxide Dismutase and Flavonoids Studied by Native Mass Spectrometry Combined with Molecular Simulations

Abstract

Misfolding and aggregation of Cu, Zn superoxide dismutase (SOD1) is implicated in the etiology of amyotrophic lateral sclerosis (ALS). The use of small molecules may stabilize the spatial structure of SOD1 dimer, thus, preventing its dissociation and aggregation. In this study, "native" mass spectrometry (MS) was used to study the noncovalent interactions between SOD1 and flavonoid compounds. MS experiments were performed on a quadruple time-of-flight (Q-ToF) mass spectrometer with an electrospray ionization (ESI) source and T-wave ion mobility. ESI-MS was used to detect the SOD1-flavonoid complexes and compare their relative binding strengths. The complement of ion mobility separation allowed comparison in the binding affinities between flavonoid isomers and provided information on the conformational changes. Molecular docking together with molecular dynamics simulations and MM/PBSA methods were applied to gain insights into the binding modes and free energies of SOD1-flavonoid complexes at the molecule level. Among all the flavonoids investigated, flavonoid glycosides preferentially bind to SOD1 than their aglycone counterparts. Naringin, one of the compounds that has the strongest binding affinity to SOD1, was subjected to further characterization. Experiment results show that the binding of naringin can stabilize SOD1 dimer and inhibit the aggregation of SOD1. Molecular simulation results suggest that naringin could reduce the dissociation of SOD1 dimers through direct interaction with the dimer interface. This developed analytical strategy could also be applied to study the interactions between SOD1 and other drug-like molecules, which may have the effect to reduce the aggregation.

Effects of aprotic solvents on the stability of metal‐free superoxide dismutase probed by native electrospray ionization–ion mobility–mass spectrometry

“…Holoenzyme SOD1 was demetaled according to previous method with slight modification. First, the stock solution of SOD1 was prepared by dialysis in a MicroFloat‐A‐Lyzer G2 against 20mM aqueous NH 4 OAc buffer at pH 3.2 containing 5mM EDTA to remove the bound metal ions.…”

Recent Developments and Applications of the MMPBSA Method

“…Many targets were studied for Alzheimer's disease including β-secretase (Koukoulitsa et al, 2016 ), angiotensin-converting enzyme (Bhavaraju et al, 2016 ), acetylcholinesterase and butyrylcholinesterase (Kurt et al, 2017 ), and inhibitors directly targeting amyloid aggregation (Berhanu and Masunov, 2015 ). Ataxin-2 protein was studied for the treatment of spinocerebellar ataxia (Sinha et al, 2017 ), superoxide dismutase for amyotrophic lateral sclerosis (Zhuang et al, 2016 ), and Niemann-Pick type Cl and C2 proteins (Poongavanam et al, 2016 ) that occur in rare neurodegenerative diseases.…”

Stabilities of superoxide dismutase and metal‐free superoxide dismutase studied by electrospray ionization ion mobility mass spectrometry

“…The metal-free SOD1 was prepared according to a previous method 14 The CIU method was used to study the conformational stability of the protein.…”