Expression and deletion analysis of the Trypanosoma brucei rhodesiense cysteine protease in Escherichia coli

E. G. Pamer, C. E. Davis, Magdalene "Maggie" So

Research output: Contribution to journalArticle

21 Citations (Scopus)

Abstract

Trypanosoma brucei, the cause of African sleeping sickness, differentiates in the mammalian bloodstream from a long, slender trypanosome into a short, stumpy trypanosome. This event is necessary for infection of the tsetse fly and maintenance of the life cycle. We have previously shown that the stumpy form contains 10- to 15-fold-greater cysteine protease activity than either the slender form or the insect midgut procyclic, and we have isolated a cDNA encoding the protease. In order to determine whether the cDNA encodes the developmentally regulated cysteine protease, we have purified the protease from trypanosomes and have made a polyclonal antiserum against it. The trypanosomal protease gene was then expressed in Escherichia coli with three different methionines within the pre- and propeptides acting as initiation sites. In each case, a protein was synthesized that was recognized by an antiserum specific for the developmentally regulated trypanosomal cysteine protease. The protein synthesized from the more upstream initiation site within the propeptide was proteolytically active. The recombinant protease and the trypanosomal enzyme were identical with respect to peptide substrates and protease inhibitors. The protein remained active when synthesized in a truncated form lacking the nine consecutive prolines and carboxy-terminus extension, indicating that the terminal 108 amino acids are not necessary for proteolytic activity.

Original languageEnglish (US)
Pages (from-to)1074-1078
Number of pages5
JournalInfection and Immunity
Volume59
Issue number3
StatePublished - 1991
Externally publishedYes

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Trypanosoma brucei rhodesiense
Cysteine Proteases
Trypanosomiasis
Peptide Hydrolases
Escherichia coli
Immune Sera
Complementary DNA
African Trypanosomiasis
Tsetse Flies
Trypanosoma brucei brucei
Proteins
Protease Inhibitors
Life Cycle Stages
Proline
Methionine
Insects
Maintenance
Amino Acids
Peptides
Enzymes

ASJC Scopus subject areas

  • Immunology

Cite this

Expression and deletion analysis of the Trypanosoma brucei rhodesiense cysteine protease in Escherichia coli. / Pamer, E. G.; Davis, C. E.; So, Magdalene "Maggie".

In: Infection and Immunity, Vol. 59, No. 3, 1991, p. 1074-1078.

Research output: Contribution to journalArticle

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