J. Ranie et al., CLONING OF THE TRIOSEPHOSPHATE ISOMERASE GENE OF PLASMODIUM-FALCIPARUM AND EXPRESSION IN ESCHERICHIA-COLI, Molecular and biochemical parasitology, 61(2), 1993, pp. 159-169
A major supply of energy in the rapidly multiplying intraerythrocytic
Plasmodium falciparum is from the glycolytic pathway. We have isolated
the cDNA and genomic clones of the glycolytic enzyme, triosephosphate
isomerase (TPI) by polymerase chain reaction (PCR). Degenerate oligon
ucleotides obtained by reverse translation of conserved polypeptide se
quences derived from TPIs of other organisms, were used to prime PCR o
n P. falciparum DNA. The P. falciparum TPI gene is interrupted by a si
ngle intron which divides the coding region into two exons. The coding
region encodes a protein of 248 amino acids which is of the same size
as TPIs from other organisms and shares 42-45% homology with other kn
own eukaryotic TPIs. On comparison with human TPI the catalytic domain
was found to be highly conserved, while significant variations occurr
ed at the other regions in the protein sequence. The P. falciparum TPI
gene was cloned into the expression vector pTrc99A and hyperexpressed
as an unfused protein in Escherichia coli. The 28-kDa protein was sho
wn to be catalytically active.