Molecular and biochemical characterization of squalene synthase from Siraitia grosvenorii
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To clone and characterize the squalene synthase from Siraitia grosvenorii (SgSQS).
The gene encoding SgSQS was cloned. SgSQS has 417 amino acid residues with an pI of 7.3. There are 32 phosphorylation sites in its sequence: S48 as well as S196 play important roles in regulation of enzyme activity. The enzyme is a monomeric protein with a cave-like active center formed by α helixes and has two transmembrane domains at its C-terminus. SgSQS mRNA expression in stem and root were about twice as much as that in leaf and peel. Full-length SgSQS with measurable catalytic activity was expressed in Escherichia coli. SgSQS activity was optimal at 37 °C and pH 7.5 respectively.
SgSQS gene was cloned, and the molecular structure and biochemical function of SgSQS were characterized.
KeywordsCloning Enzymatic properties Gene expression Sequence analysis Siraitia grosvenorii Squalene synthase
This work was supported by the National Natural Science Foundation of China (31260069) and the Training Project of the Outstanding Higher Education Teachers of Guangxi (2014) supported by the Education Department of Guanxi Zhuang Autonomous Region, China. We thank Dr. Shanhong Ling at Monash University, Melbourne, Australia, for his help in the manuscript preparation and edition.
Supplementary Fig. 1—Phylogenetic tree of the squalene synthases from Siraitia grosvenorii and other terrestrial plants.
Supplementary Fig. 2—Analysis of the tissue-specific expression of the squalene synthase from Siraitia grosvenorii using qRT-PCR.
Supplementary Fig. 3—Activity assay of the recombinant squalene synthase from Siraitia grosvenorii (SgSQS) for determination of K m and V max values.
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