S. Attucci et al., FARNESYL PYROPHOSPHATE SYNTHASE FROM WHITE LUPIN - MOLECULAR-CLONING,EXPRESSION, AND PURIFICATION OF THE EXPRESSED PROTEIN, Archives of biochemistry and biophysics, 321(2), 1995, pp. 493-500
Plants produce a variety of sesquiterpenoid compounds with diverse bio
logical functions, whose synthesis is initiated by farnesyl pyrophosph
ate synthase [EC 2.5.1.1, EC 2.5.1.10], The lack of availability of mo
lecular tools to analyze this enzyme has, thus far, prevented the stud
y of its expression and regulation in plants, A DNA fragment correspon
ding to a portion of the farnesyl pyrophosphate synthase gene was ampl
ified by the polymerase chain reaction using degenerate primers design
ed from two highly conserved domains (FLV(A/L)DD(I/M)MD and FQIQDD-DYL
D) found in eukaryotic farnesyl pyrophosphate synthase sequences, A cl
one, pS19, of a 438-bp PCR fragment was used to screen a white lupin r
oot cDNA library, Two full-length cDNA clones (pFPSI and pFPS2) were i
solated and sequenced, and one of them (pFPS2) was expressed in a bact
erial system and the enzyme protein encoded by the clone was purified,
The 1345-bp insert of pFPS2 contains a 1026-bp open reading frame, en
coding a 342-amino-acid peptide with a calculated molecular mass of 39
,310 Da. The deduced amino acid sequence of lupin farnesyl pyrophospha
te synthase pFPS2 shares 90 and 79% identity with those from Lupinus a
lbus (pFPSI) and Arabidopsis thaliana, respectively, 51% with the yeas
t enzyme, and 44% identity with those from rat and human, The overexpr
essed protein, which was purified to near homogeneity, displayed both
dimethylalkyl transferase and geranyl transferase activities, Polyclon
al antibodies raised against the purified protein immunorecognized a c
a 39-kDa protein in lupin root extracts. (C) 1995 Academic Press, Inc.