{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,8,19]],"date-time":"2026-08-19T17:48:04Z","timestamp":1787161684616,"version":"build-2736575974"},"reference-count":0,"publisher":"National Academy of Sciences","issue":"9","content-domain":{"domain":["www.pnas.org"],"crossmark-restriction":true},"short-container-title":["Proc. Natl. Acad. Sci. U.S.A."],"published-print":{"date-parts":[[1983,5]]},"abstract":"<jats:p>The catalytic role of the active site metal-water complex in horse liver alcohol dehydrogenase (alcohol:NAD+ oxidoreductase, EC 1.1.1.1) is investigated on the basis of a comparative analysis of the pH dependence of steady-state kinetic parameters of the native and active-site-specific Co2+-reconstituted enzyme and on the basis of assignment of the coordination environment of the Co2+ by electron paramagnetic resonance methods. The pH dependence of the kinetic parameters for the oxidation of benzyl alcohol reveals two ionizations (pK1 approximately equal to 6.7; pK2 approximately equal to 10.6) that govern kcat and belong to the ternary enzyme-NAD+-alcohol complex and two ionizations (pK1' approximately equal to 7.5; pK2' approximately equal to 8.9) that govern kcat\/Km and belong to the binary enzyme-NAD+ complex. The ionizations pK2 and pK2' decrease by 0.5-1 pK alpha unit upon replacement of the active site Zn2+ by Co2+. A similar metal ion dependence of pK2 and pK2' is observed for the oxidation of 2-propanol. We attribute these ionizations to a metal-bound water molecule. The zero-field splitting energy of the Co2+ in the binary enzyme-NADH complex and the ternary enzyme-NADH-CF3CH2OH complex is approximately equal to 22 cm-1, indicative of a pentacoordinate species. Binding of a water molecule to the metal ion as the fifth ligand in the ternary enzyme-NADH-CF3CH2OH complex is confirmed on the basis of magnetic interactions of H2(17)O with Co2+. The results indicate that the active site metal ion in catalytically competent ternary enzyme-coenzyme-substrate complexes is pentacoordinate and is ligated by a neutral water molecule in the physiological pH range. We suggest that the neutral metal-bound water molecule serves as the base catalyst for proton abstraction in alcohol oxidation.<\/jats:p>","DOI":"10.1073\/pnas.80.9.2584","type":"journal-article","created":{"date-parts":[[2006,5,31]],"date-time":"2006-05-31T05:21:26Z","timestamp":1149052886000},"page":"2584-2588","update-policy":"https:\/\/doi.org\/10.1073\/pnas.cm10313","source":"Crossref","is-referenced-by-count":44,"title":["Neutral metal-bound water is the base catalyst in liver alcohol dehydrogenase."],"prefix":"10.1073","volume":"80","author":[{"given":"M W","family":"Makinen","sequence":"first","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"W","family":"Maret","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"M B","family":"Yim","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"341","published-online":{"date-parts":[[1983,5]]},"container-title":["Proceedings of the National Academy of Sciences"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/pnas.org\/doi\/pdf\/10.1073\/pnas.80.9.2584","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2022,4,13]],"date-time":"2022-04-13T12:03:42Z","timestamp":1649851422000},"score":1,"resource":{"primary":{"URL":"https:\/\/pnas.org\/doi\/full\/10.1073\/pnas.80.9.2584"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[1983,5]]},"references-count":0,"journal-issue":{"issue":"9","published-print":{"date-parts":[[1983,5]]}},"alternative-id":["10.1073\/pnas.80.9.2584"],"URL":"https:\/\/doi.org\/10.1073\/pnas.80.9.2584","relation":{},"ISSN":["0027-8424","1091-6490"],"issn-type":[{"value":"0027-8424","type":"print"},{"value":"1091-6490","type":"electronic"}],"subject":[],"published":{"date-parts":[[1983,5]]},"assertion":[{"value":"1983-05-01","order":2,"name":"published","label":"Published","group":{"name":"publication_history","label":"Publication History"}}]}}