Disulfide bond involvement in the maintenance of the cryptic nature of the cross-reacting determinant of metacyclic forms of Trypanosoma congolense.
The variable surface glycoprotein (VSG) of African trypanosomes possesses a 1,2-dimyristoylglycosylphosphatidylinositol at the carboxy terminus. Cleavage of the 1,2-dimyristoylglycerol (1,2-DMG) moiety from the VSG reportedly results in a higher apparent molecular mass and an increased binding of antibodies against the "cross-reacting determinant" (CRD), a cryptic epitope present on most VSGs. Using metacyclic forms of Trypanosoma congolense, we show that the processes involved are more complex than heretofore presumed and that the removal of the 1,2-DMG moiety may not be necessary for binding of anti-CRD antibodies (RxCRD). Among other findings, we observe the following: (1) in sonicated samples of trypanosomes metabolically labeled with [3H]myristate, the binding of RxCRD on Western blots is coincident with bands containing labeled (membrane form) VSGs; (2) disulfide reduction of trypanosome sonicates suffices to promote RxCRD binding in the presence or absence of inhibitors of a glycosylphosphatidylinositol-specific phospholipase C; (3) trypanosomes directly solubilized in detergents show quantitative and qualitative differences in RxCRD binding which depend upon the detergent used and the order of addition of disulfide reducing agents. We conclude that the binding of RxCRD to T. congolense metacyclic VSGs depends upon the degree of unfolding of the molecule and is clearly a complex, multistep process in which structural changes and disulfide reduction play pivotal roles.
Fish
W R
WR
International Laboratory for Research on Animal Diseases, Nairobi, Kenya.
Muriuki
C W
CW
Muthiani
A M
AM
Grab
D J
DJ
Lonsdale-Eccles
J D
JD
D000818:Animals; D000937:Antigen-Antibody Reactions; D015153:Blotting, Western; D003429:Cross Reactions; D004220:Disulfides / Q000032:analysis; D004591:Electrophoresis, Polyacrylamide Gel; D000939:Epitopes / Q000032:analysis*; D007074:Immunoglobulin G; D008970:Molecular Weight; D019814:Myristic Acid; D009227:Myristic Acids / Q000378:metabolism; D014348:Trypanosoma congolense / Q000276:immunology*; D014643:Variant Surface Glycoproteins, Trypanosoma / Q000276:immunology
D004220:Disulfides; D000939:Epitopes; D007074:Immunoglobulin G; D009227:Myristic Acids; D014643:Variant Surface Glycoproteins, Trypanosoma; D019814:Myristic Acid