T. Imajo et al., IMMUNOCHEMICALLY DISTINCT NADP-LINKED ISOCITRATE DEHYDROGENASE ISOZYMES IN MITOCHONDRIA AND PEROXISOMES OF CANDIDA-TROPICALIS, Archives of microbiology, 168(5), 1997, pp. 389-395
Although peroxisomal localization of NADP-linked isocitrate dehydrogen
ase (Idp) was first demonstrated in Candida tropicalis, the mitochondr
ial isozyme has not been found in this yeast. Here we report that the
presence of mitochondrial Idp in the yeast was demonstrated by screeni
ng for its gene with a DNA probe containing conserved sequences of Idp
s from various organisms. The nucleotide sequence of the gene (CtIDP1)
revealed a 1,290-bp open reading frame corresponding to a 430-amino-a
cid protein with a high similarity to previously reported Idps, Overex
pression of CtIDP1 in Saccharomyces cerevisiae gave a high intracellul
ar Idp activity, and the purified recombinant Idp was shown to be a ho
modimer with a subunit molecular mass of approximately 44 kDa, differe
nt from that of peroxisomal Idp (45 kDa) previously purified from C. t
ropicalis. Western blot analysis of the subcellular fractions from ace
tate-grown C. tropicalis with polyclonal antibodies raised against the
recombinant CtIdp1 showed that the CtIdp1 in C. tropicalis was locali
zed in mitochondria but not in peroxisomes, Similar levels of CtIDP1 m
RNA and its protein product were detected in cells grown on glucose, a
cetate, and n-alkane, although a slight decrease was observed in n-alk
ane-grown cells. From these results, CtIdp1 was demonstrated to be mit
ochondrial Idp. The properties of mitochondrial Idp and peroxisomal Id
p isozymes were proven to be similar, but they were immunochemically d
istinct, suggesting the presence of another gene responsible for perox
isomal Idp in C. tropicalis.