{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2025,10,20]],"date-time":"2025-10-20T14:42:42Z","timestamp":1760971362620,"version":"build-2065373602"},"reference-count":20,"publisher":"Wiley","issue":"2","license":[{"start":{"date-parts":[[2004,1,14]],"date-time":"2004-01-14T00:00:00Z","timestamp":1074038400000},"content-version":"vor","delay-in-days":0,"URL":"http:\/\/onlinelibrary.wiley.com\/termsAndConditions#vor"}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Magnetic Reson in Chemistry"],"published-print":{"date-parts":[[2004,2]]},"abstract":"<jats:title>Abstract<\/jats:title><jats:p>The fluorinated anti\u2010psychotic drug trifluoperazine (TFP) has been shown to be a K<jats:sup>+<\/jats:sup>\u2010competitive inhibitor of gastric H<jats:sup>+<\/jats:sup>\/K<jats:sup>+<\/jats:sup>\u2010ATPase, a membrane\u2010embedded therapeutic target for peptic ulcer disease. This paper describes how variable contact time <jats:sup>19<\/jats:sup>F cross\u2010polarization magic angle spinning (VCT\u2010CP\/MAS) NMR has been used to probe the inhibitory interactions between TFP and H<jats:sup>+<\/jats:sup>\/K<jats:sup>+<\/jats:sup>\u2010ATPase in native gastric membranes. The <jats:sup>19<\/jats:sup>F CP\/MAS spectra for TFP in H<jats:sup>+<\/jats:sup>\/K<jats:sup>+<\/jats:sup>\u2010ATPase enriched (GI) gastric membranes and in control membranes containing less than 5 nmol of the protein indicated that the drug associates with the membranes independently of the presence of H<jats:sup>+<\/jats:sup>\/K<jats:sup>+<\/jats:sup>\u2010ATPase. The <jats:sup>19<\/jats:sup>F peak intensities in the VCT\u2010CP\/MAS experiment confirmed that TFP undergoes slow dissociation (<jats:italic>k<\/jats:italic><jats:sub>off<\/jats:sub> &lt; 100 s<jats:sup>\u22121<\/jats:sup>) from binding sites in GI membranes, and more rapid dissociation (<jats:italic>k<\/jats:italic><jats:sub>off<\/jats:sub> &lt; 100 s<jats:sup>\u22121<\/jats:sup>) from control membranes. The spectra showed that up to 40% of bound TFP was displaced from GI membranes by 100 m<jats:sc>M<\/jats:sc> K<jats:sup>+<\/jats:sup> and by the K<jats:sup>+<\/jats:sup>\u2010competitive inhibitor TMPIP, but TFP was not displaced from the control membranes. Hence the spectra of TFP in GI membranes represent the drug bound to the K<jats:sup>+<\/jats:sup>\u2010competitive inhibitory site of H<jats:sup>+<\/jats:sup>\/K<jats:sup>+<\/jats:sup>\u2010ATPase and to other non\u2010specific sites. The affinity of TFP for the K<jats:sup>+<\/jats:sup>\u2010competitive site (<jats:italic>K<\/jats:italic><jats:sub>D<\/jats:sub> = 4 m<jats:sc>M<\/jats:sc>) was determined from a binding curve of <jats:sup>19<\/jats:sup>F peak intensity versus TFP concentration after correction for non\u2010specific binding. The <jats:italic>K<\/jats:italic><jats:sub>D<\/jats:sub> was much higher than the IC<jats:sub>50<\/jats:sub> for ATPase inhibition (8 \u00b5<jats:sc>M<\/jats:sc>), which suggests that the substantial non\u2010specific binding of TFP to the membranes contributes to ATPase inhibition. This novel approach to probing ligand binding can be applied to a wide range of membrane\u2010embedded pharmaceutical targets, such as G\u2010protein coupled receptors and ion channels, regardless of the size of the protein or strength of binding. Copyright \u00a9 2004 John Wiley &amp; Sons, Ltd.<\/jats:p>","DOI":"10.1002\/mrc.1319","type":"journal-article","created":{"date-parts":[[2004,1,19]],"date-time":"2004-01-19T03:01:49Z","timestamp":1074481309000},"page":"204-211","source":"Crossref","is-referenced-by-count":11,"title":["Insights into the interactions between a drug and a membrane protein target by fluorine cross\u2010polarization magic angle spinning NMR"],"prefix":"10.1002","volume":"42","author":[{"given":"Martin P.","family":"Boland","sequence":"first","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"David A.","family":"Middleton","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"311","published-online":{"date-parts":[[2004,1,14]]},"reference":[{"key":"e_1_2_1_2_2","doi-asserted-by":"publisher","DOI":"10.2174\/1386207023329996"},{"key":"e_1_2_1_3_2","doi-asserted-by":"publisher","DOI":"10.1021\/ja0100120"},{"key":"e_1_2_1_4_2","doi-asserted-by":"publisher","DOI":"10.1021\/ja0264347"},{"key":"e_1_2_1_5_2","doi-asserted-by":"publisher","DOI":"10.1021\/ja992588j"},{"key":"e_1_2_1_6_2","doi-asserted-by":"publisher","DOI":"10.1021\/jm9901475"},{"key":"e_1_2_1_7_2","doi-asserted-by":"publisher","DOI":"10.1126\/science.287.5460.1960"},{"key":"e_1_2_1_8_2","doi-asserted-by":"publisher","DOI":"10.1021\/jm0109154"},{"key":"e_1_2_1_9_2","doi-asserted-by":"publisher","DOI":"10.1021\/ja020174b"},{"key":"e_1_2_1_10_2","doi-asserted-by":"publisher","DOI":"10.1021\/jm011122k"},{"key":"e_1_2_1_11_2","doi-asserted-by":"publisher","DOI":"10.1073\/pnas.91.9.3877"},{"key":"e_1_2_1_12_2","doi-asserted-by":"publisher","DOI":"10.1016\/S0006-3495(98)77722-7"},{"key":"e_1_2_1_13_2","doi-asserted-by":"publisher","DOI":"10.1016\/S0014-5793(97)00525-5"},{"key":"e_1_2_1_14_2","doi-asserted-by":"publisher","DOI":"10.1146\/annurev.ph.52.030190.001541"},{"key":"e_1_2_1_15_2","doi-asserted-by":"crossref","first-page":"5313","DOI":"10.1016\/S0021-9258(19)70788-1","volume":"255","author":"Wallmark B","year":"1980","journal-title":"J. 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