Computational analysis of a novel mutation in ETFDH gene highlights its long-range effects on the FAD-binding motif

High resolution melting (HRM) analysis and sequencing of the entire ETFDH gene revealed a novel mutation (p.Phe128Ser) and the hotspot mutation (p.Ala84Thr) from a patient with MADD. According to the predicted 3D structure of ETF:QO, the two mutations are located within the flavin adenine dinucleotide (FAD) binding domain; however, the two residues do not have direct interactions with the FAD ligand. Using molecular dynamics (MD) simulations and normal mode analysis (NMA), we found that the p.Ala84Thr and p.Phe128Ser mutations are most likely to alter the protein structure near the FAD binding site as well as disrupt the stability of the FAD binding required for the activation of ETF:QO. Intriguingly, NMA revealed that several reported disease-causing mutations in the ETF:QO protein show highly correlated motions with the FAD-binding site.Based on the present findings, we conclude that the changes made to the amino acids in ETF:QO are likely to influence the FAD-binding stability.Multiple acyl-CoA dehydrogenase deficiency (MADD), also known as glutaric aciduria type II, is an autosomal recessively inherited disorder of fatty acid metabolism [1]. In the majority of cases, the disorder is caused by a defect in either the alpha- or beta-subunit of the electron transfer flavoprotein (ETFA; OMIM 608053, ETFB; OMIM 130410) or the electron transfer flavoprotein dehydrogenase (ETFDH; OMIM 231675) genes. However, in some patients, the disorder may result from a few, currently unidentified, disruptions to riboflavin metabolism [2-4]. Because of a deficiency of ETF or ETFDH, the acyl-CoA dehydrogenases are unable to transfer the electrons generated by the dehydrogenation reactions. This results in the accumulation of various intramitochondrial acyl-CoA esters. Recently, ETFDH mutations were reported to be major causes of riboflavin-responsive MADD [5].Characteristics such as organic aciduria may be detected only during periods of illness or catabolic stress. In the majority of