{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,3,12]],"date-time":"2026-03-12T04:41:18Z","timestamp":1773290478527,"version":"3.50.1"},"reference-count":0,"publisher":"Portland Press Ltd.","issue":"2","content-domain":{"domain":["portlandpress.com"],"crossmark-restriction":true},"short-container-title":[],"published-print":{"date-parts":[[1980,11,1]]},"abstract":"<jats:p>1. Protein disulphide-isomerase (EC 5.3.4.1) and glutathione-insulin transhydrogenase (EC 1.8.4.2) were resolved by covalent chromatography. Both activities, in a partially purified preparation from bovine liver, bind covalently as mixed disulphides to activated thiopropyl-Sepharose 6B, in a new stepwise elution procedure protein disulphide-isomerase is displaced in mildly reducing conditions whereas glutathione-insulin transhydrogenase is only displaced by more extreme reducing conditions. 2. This together with evidence for partial resolution of the two activities by ion-exchange chromatography, conclusively establishes that the two activities are not alternative activities of a single bovine liver enzyme. 3. Protein disulphide-isomerase, partially purified by a published procedure, has now been further purified by covalent chromatography and ion-exchange chromatography. The final material is 560-fold purified relative to a bovine liver homogenate; it has barely detectable glutathione-insulin transhydrogenase activity. 4. The purified protein disulphide-isomerase shows a single major band on sodium dodecyl sulphate\/polyacrylamide-gel electrophoresis corresponding to a mol.wt. of 57000. 5. The purified protein disulphide-isomerase has Km values for \u2018scrambled\u2019 ribonuclease and dithiothreitol of 23 microgram\/ml and 5.4 microM respectively and has a sharp pH optimum at 7.5. The enzyme has a broad thiol-specificity, and several monothiols, at 1mM, can replace dithiothreitol. 6. The purified protein disulphide-isomerase is completely inactivated after incubation with a 2-3 fold molar excess of iodoacetate. The enzyme is also significantly inhibited by low concentrations of Cd2+ ions. These findings strongly suggest the existence of a vicinal dithiol group essential for enzyme activity. 7. When a range of thiols were used as co-substrates for protein disulphide-isomerase activity, the activities were found to co-purify quantitatively, implying the presence of a single protein disulphide-isomerase of broad thiol-specificity. Glutathione-disulphide transhydrogenase activities, assayed with a range of disulphide compounds, did not co-purify quantitatively.<\/jats:p>","DOI":"10.1042\/bj1910373","type":"journal-article","created":{"date-parts":[[2015,8,10]],"date-time":"2015-08-10T20:24:57Z","timestamp":1439238297000},"page":"373-388","update-policy":"https:\/\/doi.org\/10.1042\/crossmark_policy","source":"Crossref","is-referenced-by-count":30,"title":["Resolution of protein disulphide-isomerase and glutathione-insulin transhydrogenase activities by covalent chromatography"],"prefix":"10.1042","volume":"191","author":[{"given":"D A","family":"Hillson","sequence":"first","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"R B","family":"Freedman","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"288","container-title":["Biochemical Journal"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/portlandpress.com\/biochemj\/article-pdf\/191\/2\/373\/575268\/bj1910373.pdf","content-type":"application\/pdf","content-version":"vor","intended-application":"syndication"},{"URL":"https:\/\/portlandpress.com\/biochemj\/article-pdf\/191\/2\/373\/575268\/bj1910373.pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2021,11,25]],"date-time":"2021-11-25T14:10:26Z","timestamp":1637849426000},"score":1,"resource":{"primary":{"URL":"https:\/\/portlandpress.com\/biochemj\/article\/191\/2\/373\/17730\/Resolution-of-protein-disulphide-isomerase-and"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[1980,11,1]]},"references-count":0,"journal-issue":{"issue":"2","published-print":{"date-parts":[[1980,11,1]]}},"URL":"https:\/\/doi.org\/10.1042\/bj1910373","relation":{},"ISSN":["0264-6021"],"issn-type":[{"value":"0264-6021","type":"print"}],"subject":[],"published":{"date-parts":[[1980,11,1]]}}}