Cloning, characterization and functional expression of a cyclophilin of Entamoeba histolytica

Full-length Entamoeba histolytica cyclophilin gene (EhCyp) was isolated, ch aracterized and recombinantly expressed in bacterial cells. The deduced ami no acid sequence of EhCyp shows 60-70% identity with cyclophilins from othe r organisms and has conserved the cyclophilin signature motifs and residues involved in cyclosporin A binding. Upstream of the 501 bp open reading fra me of EhCyp, sequences resembling the putative consensus E. histolytica CE1 , CE2 and CE3 regulatory elements were found. Northern blot assays revealed a single transcript of 0.63 kb. The transcription start was determined by primer extension at position -13 relative to the initial ATG codon. Cyclosp orin A binding and peptidyl-proplyl cis-trans isomerase activities characte ristic of cyclophilin were detected in soluble extracts of E, histolytica t rophozoites and in the recombinant protein. In both cases, the isomerase ac tivity was inhibited by nanomolar concentrations of cyclosporin A. Treatmen t of cultured trophozoites with cyclosporin A decreased their proliferation with a 50% inhibition value of 1 mu g/ml and was lethal in doses over 50 m u g/ml. (C) 2000 Published by Elsevier Science B.V, All rights reserved.