{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2023,10,17]],"date-time":"2023-10-17T05:48:48Z","timestamp":1697521728924},"reference-count":0,"publisher":"Portland Press Ltd.","issue":"1","content-domain":{"domain":["portlandpress.com"],"crossmark-restriction":true},"short-container-title":[],"published-print":{"date-parts":[[1995,7,1]]},"abstract":"<jats:p>The roles of Arg-104 and Arg-225 located in the 2-kinase domain of the bifunctional enzyme 6-phosphofructo-2-kinase (PFK-2)\/fructose-2,6-bisphosphatase (FBPase-2) have been studied by site-directed mutagenesis. In recombinant rat liver PFK-2\/FBPase-2, mutation of Arg-225 to Ser increased the Km of PFK-2 for fructose-6-phosphate (Fru-6-P) 7-fold at pH 6 and decreased PFK-2 activity at suboptimal substrate concentrations between pH 6 and 9.5. The mutation had no effect on the Vmax of PFK-2 or on the Km of PFK-2 for MgATP. The mutation also increased the Vmax. of FBPase-2 4-fold without changing the Km for Fru-2,6-P2 or IC50 of Fru-6-P. These findings are in agreement with a previous study [Rider and Hue (1992) Eur. J. Biochem. 207, 967-972] on the protection by Fru-6-P of the labelling of Arg-225 by phenylglyoxal, and suggest that Arg-225 participates in Fru-6-P binding. In recombinant rat muscle PFK-2\/FBPase-2, mutation of Arg-104 to Ser increased the Km for Fru-6-P 60-fold, increased the IC50 of citrate, increased the Vmax. 1.5-3-fold at pH 8.5 and altered the pH profile of PFK-2 activity. It did not affect the Km of PFK-2 for MgATP. The mutation also decreased the Vmax. of FBPase-2 3-fold, increased the Km for Fru-2,6-P2 70-fold and increased the IC50 of Fru-6-P at least 300-fold. Although the dimeric structure was maintained in the mutant, its PFK-2 activity was more sensitive towards inactivation by guanidinium chloride than the wild-type enzyme activity. The findings indicate that Arg-104 is involved in Fru-6-P binding in the PFK-2 domain and that it might also bind citrate. Structural changes resulting from the mutation might be responsible for the changes in kinetic properties of FBPase-2.<\/jats:p>","DOI":"10.1042\/bj3090341","type":"journal-article","created":{"date-parts":[[2015,8,10]],"date-time":"2015-08-10T21:44:04Z","timestamp":1439243044000},"page":"341-346","update-policy":"http:\/\/dx.doi.org\/10.1042\/crossmark_policy","source":"Crossref","is-referenced-by-count":7,"title":["Study of the roles of Arg-104 and Arg-225 in the 2-kinase domain of 6-phosphofructo-2-kinase\/fructose-2,6-bisphosphatase by site-directed mutagenesis"],"prefix":"10.1042","volume":"309","author":[{"given":"M H","family":"Rider","sequence":"first","affiliation":[{"name":"Hormone and Metabolic Research Unit, International Institute of Cellular and Molecular Pathology, and University of Louvain Medical School, Avenue Hippocrate, 75, B-1200 Brussels, Belgium"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"K M","family":"Crepin","sequence":"additional","affiliation":[{"name":"Hormone and Metabolic Research Unit, International Institute of Cellular and Molecular Pathology, and University of Louvain Medical School, Avenue Hippocrate, 75, B-1200 Brussels, Belgium"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"M","family":"De Cloedt","sequence":"additional","affiliation":[{"name":"Hormone and Metabolic Research Unit, International Institute of Cellular and Molecular Pathology, and University of Louvain Medical School, Avenue Hippocrate, 75, B-1200 Brussels, Belgium"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"L","family":"Bertrand","sequence":"additional","affiliation":[{"name":"Hormone and Metabolic Research Unit, International Institute of Cellular and Molecular Pathology, and University of Louvain Medical School, Avenue Hippocrate, 75, B-1200 Brussels, Belgium"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"D","family":"Vertommen","sequence":"additional","affiliation":[{"name":"Hormone and Metabolic Research Unit, International Institute of Cellular and Molecular Pathology, and University of Louvain Medical School, Avenue Hippocrate, 75, B-1200 Brussels, Belgium"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"L","family":"Hue","sequence":"additional","affiliation":[{"name":"Hormone and Metabolic Research Unit, International Institute of Cellular and Molecular Pathology, and University of Louvain Medical School, Avenue Hippocrate, 75, B-1200 Brussels, Belgium"}],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"288","container-title":["Biochemical Journal"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/portlandpress.com\/biochemj\/article-pdf\/309\/1\/341\/617740\/bj3090341.pdf","content-type":"application\/pdf","content-version":"vor","intended-application":"syndication"},{"URL":"https:\/\/portlandpress.com\/biochemj\/article-pdf\/309\/1\/341\/617740\/bj3090341.pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2021,11,23]],"date-time":"2021-11-23T23:50:15Z","timestamp":1637711415000},"score":1,"resource":{"primary":{"URL":"https:\/\/portlandpress.com\/biochemj\/article\/309\/1\/341\/32301\/Study-of-the-roles-of-Arg-104-and-Arg-225-in-the-2"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[1995,7,1]]},"references-count":0,"journal-issue":{"issue":"1","published-print":{"date-parts":[[1995,7,1]]}},"URL":"https:\/\/doi.org\/10.1042\/bj3090341","relation":{},"ISSN":["0264-6021","1470-8728"],"issn-type":[{"value":"0264-6021","type":"print"},{"value":"1470-8728","type":"electronic"}],"subject":[],"published":{"date-parts":[[1995,7,1]]}}}