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A two‐domain structure for the two subunits of NAD(P)H:quinone acceptor oxidoreductase
Author(s) -
Chen Shiuan,
Deng Paulis S.K.,
Bailey Jerome M.,
Swiderek Kristine M.
Publication year - 1994
Publication title -
protein science
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 3.353
H-Index - 175
eISSN - 1469-896X
pISSN - 0961-8368
DOI - 10.1002/pro.5560030107
Subject(s) - menadione , nad+ kinase , oxidoreductase , chemistry , quinone , reductase , stereochemistry , dicoumarol , trypsin , biochemistry , enzyme , chymotrypsin , protein subunit , gene
NAD(P)H:quinone acceptor oxidoreductase (EC 1.6.99.2) (DT‐diaphorase) is a FAD‐containing reductase that catalyzes a unique 2‐electron reduction of quinones. It consists of 2 identical subunits. In this study, it was found that the carboxyl‐terminal portion of the 2 subunits can be cleaved by various proteases, whereas the amino‐terminal portion cannot. It was also found that proteolytic digestion of the enzyme can be blocked by the prosthetic group FAD, substrates NAD(P)H and menadione, and inhibitors dicoumarol and phenindione. Interestingly, chrysin and Cibacron blue, 2 additional inhibitors, cannot protect the enzyme from proteolytic digestion. The results obtained from this study indicate that the subunit of the quinone reductase has a 2‐domain structure, i.e., an aminoterminal compact domain and a carboxyl‐terminal flexible domain. A structural model of the quinone reductase is generated based on results obtained from amino‐terminal and carboxyl‐terminal protein sequence analyses and electrospray mass spectral analyses of hydrolytic products of the enzyme generated by trypsin, chymotrypsin, and Staphylococcus aureus protease. Furthermore, based on the data, it is suggested that the binding of substrates involves an interaction between 2 structural domains.