Expression and properties of diacylglycerol acyltransferase from cell-suspension cultures of oilseed rape

• Published in: Biochemical Society transactions Vol. 28; no. 6; p. 684
• Main Authors: Weselake, R J, Nykiforuk, C L, Laroche, A, Patterson, N A, Wiehler, W B, Szarka, S J, Moloney, M M, Tari, L W, Derekh, U
• Format: Journal Article
• Language: English
• Published: England 01.12.2000
• Subjects: Acyl Coenzyme A - metabolism; Acyltransferases - genetics; Acyltransferases - metabolism; Brassica - cytology; Brassica - enzymology; Cells, Cultured; Diacylglycerol O-Acyltransferase; Kinetics; Protein Biosynthesis; Recombinant Proteins - metabolism; Substrate Specificity; Time Factors; Transcription, Genetic
• ISSN: 0300-5127; 1470-8752
• DOI: 10.1042/bst0280684

The expression of diacylglycerol acyltransferase (DGAT, EC 2.3.1.20) with predicted molecular mass of 56.9. kDa (BnDGAT1) was examined using microspore-derived cell suspension cultures of oilseed rape (Brassica napus L. cv Jet Neuf). As well, a recombinant histidine-tagged N-terminal fragment of BnDGAT1 [BnDGAT1((1-116))His(6)], which was relatively hydrophilic, was partially characterized. A temporal increase in DGAT activity occurred within a 24 h period following transfer of cells from 6% (w/v) sucrose to 14% (w/v) sucrose. Western blotting indicated that the abundance of BnDGAT1 protein was closely correlated with DGAT activity. BnDGAT1 mRNA also exhibited a temporal increase within the 24 h period following transfer of cells into higher sucrose concentrations, but the transcript level was not closely associated with DGAT activity as BnDGAT1 protein. The fragment BnDGAT1(1-116)His(6) interacted with [1-(14)C]oleoyl-CoA, suggesting that the N-terminal region of BnDGAT1 may have a role in binding cellular acyl-CoA.