Extracellular release of the glycosylphosphatidylinositol (GPI)-linked Leishmania surface metalloprotease, gp63, is independent of GPI phospholipolysis -: Implications for parasite virulence

被引:90
作者
McGwire, BS
O'Connell, WA
Chang, KP
Engman, DM
机构
[1] Northwestern Univ, Sch Med, Dept Pathol, Chicago, IL 60611 USA
[2] Northwestern Univ, Sch Med, Dept Microbiol Immunol, Chicago, IL 60611 USA
[3] Univ Illinois, Coll Med, Infect Dis Sect, Chicago, IL 60612 USA
[4] Finch Univ Hlth Sci Chicago Med Sch, Dept Microbiol & Immunol, N Chicago, IL 60064 USA
关键词
D O I
10.1074/jbc.M109072200
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
The major zinc metalloprotease of Leishmania (gp63), an important determinant of parasite virulence, is attached to the parasite surface via a glycosylphosphatidylinositol anchor. Here we report the spontaneous release of proteolytically active gp63 from a number of Leishmania isolates, causing cutaneous and visceral disease. To investigate the mechanism(s) of gp63 release, we transfected a gp63-deficient variant of Leishmania amazonensis with constructs expressing gp63 and various mutants thereof. Surprisingly, approximately half of wild type gp63 was found in the culture supernatant 12 h post-synthesis. Biochemical analysis of the extracellular gp63 revealed two forms of the protein, one that is released from the cell surface, and another, that apparently is directly secreted. Release of cell surface gp63 was significantly reduced when the proteolytic activity of the protein was inactivated by site-specific mutagenesis or inhibited by zinc chelation, suggesting that release involves autoproteolysis. The extracellular gp63 does not contain a glycosylphosphatidylinositol moiety or ethanolamine, indicating that phospholipolysis is not involved in the release process. Release of gp63 is also independent of glycosylation. The finding of proteolytically active, extracellular gp63 produced by multiple Leishmania isolates suggests a potential role of the extracellular enzyme in substrate degradation relevant to their survival in both the mammalian host and the insect vector.
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页码:8802 / 8809
页数:8
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