Collapse of the native structure caused by a single amino acid exchange in human NAD(P)H:quinone oxidoreductase1. (19th September 2014)

• Record Type: Journal Article
• Authors: Lienhart, Wolf‐Dieter; Gudipati, Venugopal; Uhl, Michael K.; Binter, Alexandra; Pulido, Sergio A.; Saf, Robert; Zangger, Klaus; Gruber, Karl; Macheroux, Peter
• Is Part Of: FEBS journal. Volume 281:Number 20(2014)
• Page Start: 4691
• Page End: 4704
• Publication Date: 2014-09-19
• Subjects: Biochemistry -- Periodicals; Molecular biology -- Periodicals; Pathology, Molecular -- Periodicals; 572
• DOI: 10.1111/febs.12975
• Languages: English
• ISSNs: 1742-464X

Abstract

<abstract abstract-type="main" id="febs12975-abs-0001"> <title> <x xml:space="preserve">Abstract</x> </title> <sec id="febs12975-sec-0001" sec-type="section"> <p>Human NAD(P)H:quinone oxidoreductase 1 (NQO1) is essential for the antioxidant defense system, stabilization of tumor suppressors (e.g. p53, p33, and p73), and activation of quinone‐based chemotherapeutics. Overexpression of NQO1 in many solid tumors, coupled with its ability to convert quinone‐based chemotherapeutics into potent cytotoxic compounds, have made it a very attractive target for anticancer drugs. A naturally occurring single‐nucleotide polymorphism (C609T) leading to an amino acid exchange (P187S) has been implicated in the development of various cancers and poor survival rates following anthracyclin‐based adjuvant chemotherapy. Despite its importance for cancer prediction and therapy, the exact molecular basis for the loss of function in NQO1 P187S is currently unknown. Therefore, we solved the crystal structure of NQO1 P187S. Surprisingly, this structure is almost identical to NQO1. Employing a combination of NMR spectroscopy and limited proteolysis experiments, we demonstrated that the single amino acid exchange destabilized interactions between the core and C‐terminus, leading to depopulation of the native structure in solution. This collapse of the native structure diminished cofactor affinity and led to a less competent FAD‐binding pocket, thus severely compromising the catalytic capacity of the variant protein. Hence, our findings provide a rationale for the loss of function in NQO1 P187S with a frequently occurring single‐nucleotide polymorphism.</p> </sec> <sec id="febs12975-sec-0002" sec-type="section"> <title>Database</title> <p>Structural data are available in the Protein Data Bank under the accession numbers <ext-link ext-link-type="uri" xlink:href="http://www.rcsb.org/pdb/search/structidSearch.do?structureId=4cet" xlink:type="simple" xmlns:xlink="http://www.w3.org/1999/xlink">4cet</ext-link> (P187S variant with dicoumarol) and <ext-link ext-link-type="uri" xlink:href="http://www.rcsb.org/pdb/search/structidSearch.do?structureId=4cf6" xlink:type="simple" xmlns:xlink="http://www.w3.org/1999/xlink">4cf6</ext-link> (P187S variant with Cibacron blue).</p> </sec> <sec id="febs12975-sec-0003" sec-type="section"> <title>Structured digital abstract</title> <p> <list id="febs12975-list-0001" list-type="bullet"> <list-item> <p> <ext-link ext-link-type="uri" xlink:href="http://www.uniprot.org/uniprot/P15559" xlink:type="simple" xmlns:xlink="http://www.w3.org/1999/xlink">NQO1 P187S</ext-link> and <ext-link ext-link-type="uri" xlink:href="http://www.uniprot.org/uniprot/P15559" xlink:type="simple" xmlns:xlink="http://www.w3.org/1999/xlink">NQO1 P187S</ext-link> <ext-link ext-link-type="uri" xlink:href="http://www.ebi.ac.uk/ontology-lookup/?termId=MI:0407" xlink:type="simple" xmlns:xlink="http://www.w3.org/1999/xlink">bind</ext-link> by <ext-link ext-link-type="uri" xlink:href="http://www.ebi.ac.uk/ontology-lookup/?termId=MI:0077" xlink:type="simple" xmlns:xlink="http://www.w3.org/1999/xlink">nuclear magnetic resonance</ext-link> (<ext-link ext-link-type="uri" xlink:href="http://www.ebi.ac.uk/intact/interaction/EBI-9685543" xlink:type="simple" xmlns:xlink="http://www.w3.org/1999/xlink">View interaction</ext-link>)</p> </list-item> <list-item> <p> <ext-link ext-link-type="uri" xlink:href="http://www.uniprot.org/uniprot/P15559" xlink:type="simple" xmlns:xlink="http://www.w3.org/1999/xlink">NQO1 P187S</ext-link> and <ext-link ext-link-type="uri" xlink:href="http://www.uniprot.org/uniprot/P15559" xlink:type="simple" xmlns:xlink="http://www.w3.org/1999/xlink">NQO1 P187S</ext-link> <ext-link ext-link-type="uri" xlink:href="http://www.ebi.ac.uk/ontology-lookup/?termId=MI:0407" xlink:type="simple" xmlns:xlink="http://www.w3.org/1999/xlink">bind</ext-link> by <ext-link ext-link-type="uri" xlink:href="http://www.ebi.ac.uk/ontology-lookup/?termId=MI:0114" xlink:type="simple" xmlns:xlink="http://www.w3.org/1999/xlink">x-ray crystallography</ext-link> (<ext-link ext-link-type="uri" xlink:href="http://www.ebi.ac.uk/intact/interaction/EBI-9685561" xlink:type="simple" xmlns:xlink="http://www.w3.org/1999/xlink">1</ext-link>, <ext-link ext-link-type="uri" xlink:href="http://www.ebi.ac.uk/intact/interaction/EBI-9685555" xlink:type="simple" xmlns:xlink="http://www.w3.org/1999/xlink">2</ext-link>)</p> </list-item> <list-item> <p> <ext-link ext-link-type="uri" xlink:href="http://www.uniprot.org/uniprot/P15559" xlink:type="simple" xmlns:xlink="http://www.w3.org/1999/xlink">NQO1</ext-link> and <ext-link ext-link-type="uri" xlink:href="http://www.uniprot.org/uniprot/P15559" xlink:type="simple" xmlns:xlink="http://www.w3.org/1999/xlink">NQO1</ext-link> <ext-link ext-link-type="uri" xlink:href="http://www.ebi.ac.uk/ontology-lookup/?termId=MI:0407" xlink:type="simple" xmlns:xlink="http://www.w3.org/1999/xlink">bind</ext-link> by <ext-link ext-link-type="uri" xlink:href="http://www.ebi.ac.uk/ontology-lookup/?termId=MI:0071" xlink:type="simple" xmlns:xlink="http://www.w3.org/1999/xlink">molecular sieving</ext-link> (<ext-link ext-link-type="uri" xlink:href="http://www.ebi.ac.uk/intact/interaction/EBI-9685585" xlink:type="simple" xmlns:xlink="http://www.w3.org/1999/xlink">1</ext-link>, <ext-link ext-link-type="uri" xlink:href="http://www.ebi.ac.uk/intact/interaction/EBI-9685575" xlink:type="simple" xmlns:xlink="http://www.w3.org/1999/xlink">2</ext-link>)</p> </list-item> </list> </p> </sec> </abstract>