Intramolecular mapping of Plasmodium falciparum P126 proteolytic fragments by N-terminal amino acid sequencing.
Protein P126 (also called P140, P113, SERA, SERP1) is a major parasitophorous vacuole antigen of Plasmodium falciparum. This protein is processed upon merozoite release into 2 fragments of 73 kDa (P73) and 50 kDa (P50), which are found in the culture medium. P73 is composed of 2 polypeptides of 47 and 18 kDa linked by disulfide bridges. In the presence of leupeptin, an inhibitor of serine and cysteine proteases which inhibits merozoite release, a 56-kDa intermediate product (P56) is recovered in the culture medium instead of P50. In order to map these proteolytic fragments on the 126-kDa precursor, we purified them from Plasmodium falciparum culture medium by immunoadsorption, SDS-electrophoresis and Western blotting on PVDF membrane and determined the N termini of P126, P73 (P47 and P18), P50 and P56. Comparison of these sequences with the amino acid sequence deduced from the P126 gene allowed the mapping of the different fragments on the precursor. P47 was at the N-terminal and P18 at the C-terminal end of P126. P56 and P50 had the same N-termini and were located in the middle of P126. This latter result indicates that the proteolysis of P56-P50 occurs at the C-terminus of P56. The peptide bonds cleaved by leupeptin-insensitive activities are Glu-Thr and Gln-Asp; C-terminal sequencing of P50 will be needed to identify the leupeptin-sensitive cleavage site.
Debrabant
A
A
Unité 42 INSERM, Villeneuve D'Ascq, France.
Maes
P
P
Delplace
P
P
Dubremetz
J F
JF
Tartar
A
A
Camus
D
D
D000595:Amino Acid Sequence; D000818:Animals; D000953:Antigens, Protozoan / Q000235:genetics; D010450:Endopeptidases / Q000378:metabolism; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000235:genetics / Q000302:isolation & purification; D010449:Peptide Mapping; D010963:Plasmodium falciparum / Q000235:genetics* / Q000276:immunology / Q000378:metabolism; D011499:Protein Processing, Post-Translational; D015800:Protozoan Proteins / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism
D000953:Antigens, Protozoan; D010446:Peptide Fragments; D015800:Protozoan Proteins; D010450:Endopeptidases