Cloning and kinetic characterization of Arabidopsis thaliana solanesyl diphosphate synthase

被引:53
作者
Hirooka, K [1 ]
Bamba, T [1 ]
Fukusaki, EI [1 ]
Kobayashi, A [1 ]
机构
[1] Osaka Univ, Grad Sch Engn, Dept Biotechnol, Suita, Osaka 5650871, Japan
关键词
isoprenoid; nonaprenyl diphosphate; plastoquinone; prenyltransferase; ubiquinone;
D O I
10.1042/BJ20021311
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
trans-Long-chain prenyl diphosphate synthases catalyse the sequential condensation of isopentenyl diphosphate (C-5) units with allylic diphosphate to produce the C-30-C-50 prenyl diphosphates, which are precursors of the side chains of prenyl-quinones. Based on the relationship between product specificity and the region around the first aspartate-rich motif in trans-prenyl diphosphate synthases characterized so far, we have isolated the cDNA for a member of trans-long-chain prenyl diphosphate synthases from Arabidopsis thaliana. The cDNA was heterologously expressed in Escherichia coli, and the recombinant His(6)-tagged protein was purified and characterized. Product analysis revealed that the cDNA encodes solanesyl diphosphate (C-45) synthase (At-SPS). At-SPS utilized farnesyl diphosphate (FPP; C-15) and geranylgeranyl diphosphate (GGPP; C-20), but did not accept either the C-5 or the C-10 allylic diphosphate as a primer substrate. The Michaelis constants for FPP and GGPP were 5.73 muM and 1.61 muM respectively. We also performed an analysis of the side chains of prenylquinones extracted from the A. thaliana plant, and showed that its major prenylquinones, i.e. plastoquinone and ubiquinone, contain the C-45 prenyl moiety. This suggests that At-SPS might be devoted to the biosynthesis of either or both of the prenylquinone side chains. This is the first established trans-long-chain prenyl diphosphate synthase from a multicellular organism.
引用
收藏
页码:679 / 686
页数:8
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