Cloning and characterization of a Plasmodium falciparum cyclophilin gene that is stage-specifically expressed.
An immunosuppressive agent, cyclosporin A (CsA), has antimalarial activity in several Plasmodium species. Cyclophilins of several species including Plasmodium falciparum exhibit peptidyl-prolyl cis-trans isomerase activity which is inhibited by CsA. A gene encoding P. falciparum cyclophilin (PFCyP) was cloned and characterized. This gene has the entire coding sequence for the mature protein plus a 39-amino-acid-long N-terminal extension. Most of the amino acids predicted to be involved in the peptidyl-prolyl cis-trans isomerase activity and CsA binding are present in the cloned gene. The PFCyP also has the single highly conserved tryptophan residue that is a major determinant in the inhibition of PPIase activity by CsA. The PFCyP coding sequence with or without the N-terminal amino-acid extension was used to construct recombinant expression vectors which were transformed into E. coli. Both vectors produced enzymatically active mature PFCyP proteins that were sensitive to CsA. Northern blot analysis of RNA isolated from the synchronized parasite cultures verified the expression of PFCyP in all erythrocytic stages of the parasite, but at variable levels. The highest level of expression was observed in ring-stage parasites, a stage shown to be more susceptible to CsA. Inhibition of P. falciparum growth in vitro by CsA was re-evaluated for chloroquine-sensitive and chloroquine-resistant strains of the parasite. Essentially, there was no difference between the two strains for the concentration of CsA required to yield 50% inhibition in 48 h of exposure (0.25-0.4 microM).
Reddy
G R
GR
Department of Infectious Diseases, College of Veterinary Medicine, University of Florida, Gainesville 32611-0880, USA.
D000591:Amino Acid Isomerases / Q000037:antagonists & inhibitors / Q000235:genetics*; D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002352:Carrier Proteins / Q000037:antagonists & inhibitors / Q000235:genetics*; D003001:Cloning, Molecular; D016572:Cyclosporine / Q000494:pharmacology; D017931:DNA Primers / Q000235:genetics; D018076:DNA, Complementary / Q000235:genetics; D016054:DNA, Protozoan / Q000235:genetics; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004912:Erythrocytes / Q000469:parasitology; D004926:Escherichia coli / Q000235:genetics; D018507:Gene Expression Regulation, Developmental; D017125:Genes, Protozoan; D006801:Humans; D008969:Molecular Sequence Data; D019696:Peptidylprolyl Isomerase; D010963:Plasmodium falciparum / Q000187:drug effects / Q000235:genetics*; D016053:RNA, Protozoan / Q000235:genetics; D017386:Sequence Homology, Amino Acid
D002352:Carrier Proteins; D017931:DNA Primers; D018076:DNA, Complementary; D016054:DNA, Protozoan; D004791:Enzyme Inhibitors; D016053:RNA, Protozoan; D016572:Cyclosporine; D000591:Amino Acid Isomerases; D019696:Peptidylprolyl Isomerase