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Purification of an UDP‐glucose:flavone, 7‐ O ‐glucosyltransferase, from Silene latifolia using a specific interaction between the enzyme and phenyl‐sepharose
Author(s) -
Vellekoop P.,
Lugones L.,
van Brederode J.
Publication year - 1993
Publication title -
febs letters
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.593
H-Index - 257
eISSN - 1873-3468
pISSN - 0014-5793
DOI - 10.1016/0014-5793(93)80914-g
Subject(s) - chromatography , chemistry , sepharose , glucosyltransferase , affinity chromatography , enzyme , size exclusion chromatography , caryophyllaceae , isovitexin , flavonoid , elution , hydrophilic interaction chromatography , column chromatography , biochemistry , high performance liquid chromatography , biology , ecology , antioxidant , vitexin
An UDP‐glucose:flavonoid, 7‐ O ‐glucosyltransferase, from Silene latifolia was isolated from petals and purified 450‐fold using a combination of gel‐filtration, affinity chromatography and anion‐exchange chromatography. Affinity chromatography on a phenyl‐Sepharose CL‐4B column in combination with elution with the substrate, isovitexin (6‐ C ‐glucosylapigenin), was an especially effective purification step. A purification factor between 10 and 20 could be reached using this column. A possible mechanism for the specific interaction of the enzyme with the phenyl‐Sepharose will be discussed. This method of purification may also be applicable to other enzymes which use aromatic compounds as substrates. On a SDS‐PAGE gel a band of 54 kDa, which co‐purified with enzyme activity, could be detected in the purest fraction.