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These proteins contribute indirectly to the establishment of the transmembrane difference of electrochemical potential by catalyzing the reduction of quinone by oxidation of NAD(P)H. NDH-2s are widespread enzymes being present in the three domains of life. In this work, we explored the catalytic mechanism of NDH-2 by investigating the common elements of all NDH-2s, based on the rationale that conservation of such elements reflects their structural\/functional importance. We observed conserved sequence motifs and structural elements among 1762 NDH-2s. We identified two proton pathways possibly involved in the protonation of the quinone. Our results led us to propose the first catalytic mechanism for NDH-2 family, in which a conserved glutamate residue, E<jats:sub>172<\/jats:sub> (in NDH-2 from <jats:italic>Staphylococcus aureus<\/jats:italic>) plays a key role in proton transfer to the quinone pocket. This catalytic mechanism may also be extended to the other members of the two-Dinucleotide Binding Domains Flavoprotein (tDBDF) superfamily, such as sulfide:quinone oxidoreductases.<\/jats:p>","DOI":"10.1038\/srep42303","type":"journal-article","created":{"date-parts":[[2017,2,9]],"date-time":"2017-02-09T10:25:40Z","timestamp":1486635940000},"update-policy":"https:\/\/doi.org\/10.1007\/springer_crossmark_policy","source":"Crossref","is-referenced-by-count":36,"title":["Structural and Functional insights into the catalytic mechanism of the Type II NADH:quinone oxidoreductase family"],"prefix":"10.1038","volume":"7","author":[{"given":"Bruno C.","family":"Marreiros","sequence":"first","affiliation":[]},{"given":"Filipa V.","family":"Sena","sequence":"additional","affiliation":[]},{"given":"Filipe M.","family":"Sousa","sequence":"additional","affiliation":[]},{"given":"A. 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