Protein geranylgeranyltransferase type I (PGGT-I) and protein farnesyltransferase (PFT) o
ccur in many eukaryoti
c cells. Both
consist of two subunits, the
common α subunit and a distin
ct β subunit. In the gene database of protozoa
Trypanosoma cruzi, the
causative agent of Chagas’ disease, a putative protein that
consists of 401 amino a
cids with
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cid sequen
ce identity to the PGGT-I β of other spe
cies was identified,
cloned, and
chara
cterized. Multiple sequen
ce alignments show that the
T. cruzi ortholog
contains all three of the zin
c-binding residues and several residues uniquely
conserved in the β subunit of PGGT-I. Co-expression of this protein and the α subunit of
T. cruzi PFT in
Sf9 inse
ct
cells yielded a dimeri
c protein that forms a tight
complex sele
ctively with [
3H]geranylgeranyl pyrophosphate, indi
cating a key
chara
cteristi
c of a fun
ctional PGGT-I. Re
combinant
T. cruzi PGGT-I ortholog showed geranylgeranyltransferase a
ctivity with distin
ct spe
cifi
city toward the C-terminal CaaX motif of protein substrates
compared to that of the mammalian PGGT-I and
T. cruzi PFT. Most of the CaaX-
containing proteins with X = Leu are good substrates of
T. cruzi PGGT-I, and those with X = Met are substrates for both
T. cruzi PFT and PGGT-I, whereas unlike mammalian PGGT-I, those with X = Phe are poor substrates for
T. cruzi PGGT-I. Several
candidates for
T. cruzi PGGT-I or PFT substrates
containing the C-terminal CaaX motif are found in the
T. cruzi gene database. Among five C-terminal peptides of those tested, a peptide of a Ras-like protein ending with CVLL was sele
ctively geranylgeranylated by
T. cruzi PGGT-I. Other peptides with CTQQ (T
cj2 DNAJ protein), CAVM (T
cPRL-1 protein tyrosine phosphatase), CHFM (a small GTPase like protein), and CQLF (T
cRho1 GTPase) were spe
cifi
c substrates for
T. cruzi PFT but not for PGGT-I. The mRNA and protein of the
T. cruzi PGGT-I β ortholog were dete
cted in three life-
cy
cle stages of
T. cruzi. Cytosol fra
ctions from trypomastigotes (infe
ctious mammalian stage) and epimastigotes (inse
ct stage) were shown to
contain levels of PGGT-I a
ctivity that are
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ctivity. The CaaX mimeti
cs known as PGGT-I inhibitors show very low poten
cy against
T. cruzi PGGT-I
compared to the mammalian enzyme, suggesting the potential to develop sele
ctive inhibitors against the parasite enzyme.