A cathepsin B-like protease is required for host protein degradation in Trypanosoma brucei.
Identification and analysis of Clan CA (papain) cysteine proteases in primitive protozoa and metazoa have suggested that this enzyme family is more diverse and biologically important than originally thought. The protozoan parasite Trypanosoma brucei is the etiological agent of African sleeping sickness. The cysteine protease activity of this organism is a validated drug target as first recognized by the killing of the parasite with the diazomethane inhibitor Z-Phe-Ala-CHN(2) (where Z is benzyloxycarbonyl). Whereas the presumed target of this inhibitor was rhodesain (also brucipain, trypanopain), the major cathepsin L-like cysteine protease of T. brucei, genomic analysis has now identified tbcatB, a cathepsin B-like cysteine protease as a possible inhibitor target. The mRNA of tbcatB is more abundantly expressed in the bloodstream versus the procyclic form of the parasite. Induction of RNA interference against rhodesain did not result in an abnormal phenotype in cultured T. brucei. However, induction of RNA interference against tbcatB led to enlargement of the endosome, accumulation of fluorescein isothiocyanate-transferrin, defective cytokinesis after completion of mitosis, and ultimately the death of cultured parasites. Therefore, tbcatB, but not rhodesain, is essential for T. brucei survival in culture and is the most likely target of the diazomethane protease inhibitor Z-Phe-Ala-CHN(2) in T. brucei.
Department of Pathology Tropical Disease Research Unit, University of California, San Francisco, California 94143, USA. zmackey@itsa.ucsf.edu
Zachary B
ZB
Mackey
Theresa C
TC
O'Brien
Doron C
DC
Greenbaum
Rebecca B
RB
Blank
James H
JH
McKerrow
D000595:Amino Acid Sequence; D000818:Animals; D002401:Cathepsin B / Q000235:genetics / Q000378:metabolism*; D048430:Cell Shape; D003546:Cysteine Endopeptidases / Q000235:genetics / Q000378:metabolism*; D015853:Cysteine Proteinase Inhibitors / Q000378:metabolism; D003978:Diazomethane / Q000737:chemistry / Q000378:metabolism; D006801:Humans; D008969:Molecular Sequence Data; D015800:Protozoan Proteins / Q000235:genetics / Q000378:metabolism*; D034622:RNA Interference; D016415:Sequence Alignment; D014168:Transferrin / Q000378:metabolism; D014346:Trypanosoma brucei brucei / Q000166:cytology / Q000201:enzymology* / Q000235:genetics; D014353:Trypanosomiasis, African / Q000469:parasitology
D015853:Cysteine Proteinase Inhibitors; D015800:Protozoan Proteins; D014168:Transferrin; D003978:Diazomethane; D003546:Cysteine Endopeptidases; C446033:rhodesain; D002401:Cathepsin B