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Immobilized uroporphyrinogen I synthetase from Rhodopseudomonas palustris
Author(s) -
Kotler ML,
Juknat AA,
Batlle AM
Publication year - 1990
Publication title -
biotechnology and applied biochemistry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.468
H-Index - 70
eISSN - 1470-8744
pISSN - 0885-4513
DOI - 10.1111/j.1470-8744.1990.tb00097.x
Subject(s) - sepharose , rhodopseudomonas palustris , chemistry , enzyme , substrate (aquarium) , chromatography , specific activity , yield (engineering) , biochemistry , incubation , affinity chromatography , biology , bacteria , ecology , genetics , materials science , metallurgy
Rhodopseudomonas palustris uroporphyrinogen I synthetase (URO‐S) has been chemically attached to Sepharose 4B and some of its properties have been studied. When 7–8 mg protein/ml activated Sepharose was used, immobilized URO‐S retained 45% of the activity of the original soluble preparation, with a coupling yield of 66% after a period of 15 h. Optimal incubation conditions for the activity of gel‐enzyme were determined. Unlike the soluble enzyme, the Sepharose‐bound URO‐S showed a biphasic substrate saturation curve, indicating that a protein conformational change had occurred during the process of immobilization. Immobilized URO‐S stored at 4 degrees C for 35 days retained 90% of activity and when repeatedly used, up to 5 times, retained 48% of the original activity. Attachment of URO‐S to Sepharose led to an enhanced thermal stability.