S-Myristoylation of a Glycosylphosphatidylinositol-specific Phospholipase C inTrypanosoma brucei*
Abstract
Covalent modification with lipid can target cytosolic proteins to biological membranes. With intrinsic membrane proteins, the role of acylation can be elusive. Herein, we describe covalent lipid modification of an integral membrane glycosylphosphatidylinositol-specific phospholipase C (GPI-PLC) from the kinetoplastid Trypanosoma brucei. Myristic acid was detected on cysteine residue(s) (i.e. thiomyristoylation). Thiomyristoylation occurred both co- and post-translationally. Acylated GPI-PLC was active against variant surface glycoprotein (VSG). The half-life of fatty acid on GPI-PLC was 45 min, signifying the dynamic nature of the modification. Deacylation in vitro decreased activity of GPI-PLC 18–30-fold. Thioacylation, from kinetic analysis, activated GPI-PLC by accelerating the conversion of a GPI-PLC·VSG complex to product. Reversible thioacylation is a novel mechanism for regulating the activity of a phospholipase C.
Footnotes
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↵* This work was supported by National Institutes of Health Grant AI33383 and by a Burroughs Wellcome Fund New Investigator Award in Molecular Parasitology (to K. M.-W.).The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
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↵‡ To whom correspondence should be addressed: Dept. of Cellular Biology, the University of Georgia, 724 Biological Sciences, Athens, GA 30602. Tel.: 706-542-3355; Fax: 706-542-4271; E-mail:mensawil{at}cb.uga.edu.
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↵2 K. Mensa-Wilmot and R. Howard, unpublished observations.
- Abbreviations:
- VSG
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variant surface glycoprotein from T. brucei
- CHX
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cycloheximide
- FAMES
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fatty acid methyl esters
- GPI
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glycosylphosphatidylinositol
- GPI-PLC
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glycosylphosphatidylinositol-specific phospholipase C
- PBS
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phosphate-buffered saline
- RP-HPTLC
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reverse phase-high performance thin layer chromatography
- PAGE
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polyacrylamide gel electrophoresis
- mfVSG
-
membrane form VSG
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- Received October 28, 1998.
- Revision received December 9, 1998.
- The American Society for Biochemistry and Molecular Biology, Inc.
