Expression of recombinant Toxoplasma gondii P24.
The gene encoding Toxoplasma gondii P24 has been reported previously. To determine the function of P24 against immune systems in the near future, we prepared recombinant P24 antigens using Escherichia coli, insect cells infected with recombinant baculovirus and mammalian cells infected with recombinant vaccinia virus. The P24 antigens derived from E. coli, insect cells and mammalian cells were detected with mouse immune sera against P24 or T. gondii homogenates by Western blot analysis; these corresponded to the authentic P24 and secreted into the supernatants of the insect and mammalian cell cultures. These proteins were not effected by tunicamycin treatment in cultured cells, indicating that recombinant P24 did not contain N-linked sugars. Recombinant P24 was separated by two-dimensional electrophoresis and analyzed by Western blotting. From these results, P24 was acidic protein and had identical isoelectric point with the authentic P24.
Tanaka
T
T
The Research Center for Protozoan Molecular Immunology, Obihiro University of Agriculture and Veterinary Medicine, Hokkaido, Japan.
Xuan
X
X
Kato
M
M
Nagasawa
H
H
Fujisaki
K
K
Mikami
T
T
Suzuki
N
N
D000818:Animals; D000900:Anti-Bacterial Agents / Q000737:chemistry; D000953:Antigens, Protozoan / Q000737:chemistry / Q000235:genetics* / Q000276:immunology; D015153:Blotting, Western; D002460:Cell Line; D003001:Cloning, Molecular; D017931:DNA Primers / Q000737:chemistry; D016054:DNA, Protozoan / Q000737:chemistry; D004587:Electrophoresis, Agar Gel; D005260:Female; D019084:Fluorescent Antibody Technique, Indirect; D005786:Gene Expression Regulation; D051379:Mice; D008807:Mice, Inbred BALB C; D008969:Molecular Sequence Data; D015800:Protozoan Proteins / Q000737:chemistry / Q000235:genetics* / Q000276:immunology; D016053:RNA, Protozoan / Q000737:chemistry / Q000302:isolation & purification; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000276:immunology; D020133:Reverse Transcriptase Polymerase Chain Reaction; D018411:Spodoptera; D014122:Toxoplasma / Q000737:chemistry / Q000235:genetics* / Q000276:immunology; D014124:Toxoplasmosis, Animal / Q000276:immunology / Q000469:parasitology*; D014162:Transfection; D014415:Tunicamycin / Q000737:chemistry
D000900:Anti-Bacterial Agents; D000953:Antigens, Protozoan; D017931:DNA Primers; D016054:DNA, Protozoan; D015800:Protozoan Proteins; D016053:RNA, Protozoan; D011994:Recombinant Proteins; D014415:Tunicamycin