Isolation of a cDNA encoding the alpha-subunit of CAAX-prenyltransferases from Catharanthus roseus and the expression of the active recombinant protein farnesyltransferase.
Crfta/ggt_Ia (AF525030), a cDNA encoding the ?-subunit of the two types of CaaX-prenyltransferase (CaaX-PTase), i.e. protein farnesyltransferase (PFT) and type I protein geranylgeranyltransferase, was cloned from Catharanthus roseus via a PCR strategy. Crfta/ggt_Ia is 1381-bp long and bears a 999-bp open reading frame encoding a protein of 332 residues (FTA) that shares 66% identity with its Lycopersicon esculentum orthologue. Southern blot analysis revealed that FTA is encoded by a single gene copy per haploid genome. Co-expression of Crfta/ggt_Ia and Crftb encoding the beta-subunit of PFT yielded purified active recombinant PFT. This enzyme is able to prenylate proteins from C. roseus, and could be used as a potent tool for prenylated protein identification.
Courdavault
Vincent
V
Université François-Rabelais de Tours, EA 2106 Biomolécules et Biotechnologies Végétales, UFR Sciences et Techniques, Parc de Grandmont, 37200 Tours, France. courdavault@univ-tours.fr
Burlat
Vincent
V
St-Pierre
Benoit
B
Gantet
Pascal
P
Giglioli-Guivarc'h
Nathalie
N
D000595:Amino Acid Sequence; D015139:Blotting, Southern; D030008:Catharanthus / Q000201:enzymology* / Q000235:genetics*; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000235:genetics* / Q000302:isolation & purification; D004122:Dimethylallyltranstransferase / Q000737:chemistry* / Q000235:genetics*; D051231:Farnesyltranstransferase / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D008969:Molecular Sequence Data; D018515:Plant Leaves; D010940:Plant Proteins; D017035:Pravastatin / Q000494:pharmacology; D017368:Protein Prenylation / Q000187:drug effects; D021122:Protein Subunits / Q000737:chemistry / Q000235:genetics; D011994:Recombinant Proteins; D016415:Sequence Alignment
D018076:DNA, Complementary; D010940:Plant Proteins; D021122:Protein Subunits; D011994:Recombinant Proteins; D004122:Dimethylallyltranstransferase; D051231:Farnesyltranstransferase; D017035:Pravastatin