Molecular characterization of a Leishmania donovanii cDNA clone with similarity to human 20S proteasome a-type subunit.
Using plasma from patients infected or previously infected with Leishmania donovanii, we isolated a L. donovanii cDNA clone with similarity to the proteasome a-type subunit from humans and other eukaryotes. The cDNA clone, designated LePa, was DNA sequenced and Northern blot analysis of L. donovanii poly(A(+))mRNA indicated the isolation of a full length cDNA clone with a transcript size of 1.9 kb. The expressed recombinant LePa fusion protein induced proliferation of peripheral blood mononuclear cells in one out of seven patients who had suffered from visceral leishmaniasis. Plasma from 16 out of 25 patients with visceral leishmaniasis and four out of 18 patients with cutaneous leishmaniasis contained IgG antibodies which reacted with the purified LePa fusion protein as evaluated in an ELISA. The LePa DNA sequence was inserted into an eukaryotic expression vector and Balb/c mice were vaccinated. DNA vaccination of Balb/c mice with LePa generated an initial significant reduction in lesion size after challenge.
Christensen
C B
CB
Centre for Medical Parasitology, Institute of Medical Microbiology and Immunology, University of Copenhagen, Blegdamsvej 3, 2200, Copenhagen, Denmark.
Jørgensen
L
L
Jensen
A T
AT
Gasim
S
S
Chen
M
M
Kharazmi
A
A
Theander
T G
TG
Andresen
K
K
D000818:Animals; D000913:Antibodies, Protozoan / Q000097:blood; D001483:Base Sequence; D015152:Blotting, Northern; D015153:Blotting, Western; D003001:Cloning, Molecular; D003546:Cysteine Endopeptidases / Q000235:genetics*; D018076:DNA, Complementary / Q000235:genetics*; D004797:Enzyme-Linked Immunosorbent Assay; D006801:Humans; D007893:Leishmania donovani / Q000235:genetics* / Q000276:immunology / Q000302:isolation & purification; D007898:Leishmaniasis, Visceral / Q000276:immunology / Q000469:parasitology / Q000517:prevention & control*; D008213:Lymphocyte Activation / Q000187:drug effects; D051379:Mice; D008807:Mice, Inbred BALB C; D008969:Molecular Sequence Data; D009097:Multienzyme Complexes / Q000235:genetics*; D046988:Proteasome Endopeptidase Complex; D011993:Recombinant Fusion Proteins / Q000276:immunology / Q000494:pharmacology; D017422:Sequence Analysis, DNA; D017386:Sequence Homology, Amino Acid; D019444:Vaccines, DNA / Q000276:immunology
D000913:Antibodies, Protozoan; D018076:DNA, Complementary; D009097:Multienzyme Complexes; D011993:Recombinant Fusion Proteins; D019444:Vaccines, DNA; D003546:Cysteine Endopeptidases; D046988:Proteasome Endopeptidase Complex