Biosynthesis of Leishmania lipophosphoglycan: solubilization and partial characterization of the initiating mannosylphosphoryltransferase.
Lipophosphoglycan (LPG) is the predominant surface glycoconjugate of Leishmania promastigotes and consists of a capped polymer of Gal(beta1,4)Man(alpha1)-PO4 repeating units attached through a glycan core to a phosphatidylinositol anchor. We have solubilized the mannosylphosphoryltransferase from L. donovani promastigotes that initiates repeating unit synthesis using beta-dodecylmaltoside and other nonionic detergents with long alkyl chains. The detergent-solubilized enzyme, in the presence of GDP-Man and Mn2+, transferred Man(alpha1)-PO4 to two exogenous acceptor substrates: the glycan core from LPG and stachyose, a tetrasaccharide terminating in the same Gal(alpha1,6)Gal(alpha) disaccharide as glycan core. The activity is saturable with respect to GDP-Man, but not with respect to stachyose, suggesting that more than Gal(alpha1,6)Gal(alpha) is required in the acceptor substrate for optimal activity. In contrast to promastigotes, Leishmania amastigotes express lower levels of LPG by downregulating the addition of the repeating units. We compared the relative activity of the initiating mannosylphosphoryltransferase in microsomal fractions from axenic amastigotes and its promastigote counterpart, using stachyose as the acceptor substrate. The promastigote membranes were 3-fold more active relative to the amastigote membranes. These results provide evidence that the initiating mannosylphosphoryltransferase is developmentally regulated during the life-cycle of the Leishmania parasite.
Mengeling
B J
BJ
Department of Biochemistry, University of Kentucky Medical Center, Lexington 40536, USA.
Zilberstein
D
D
Turco
S J
SJ
D000818:Animals; D002240:Carbohydrate Sequence; D006028:Glycosphingolipids / Q000096:biosynthesis*; D007893:Leishmania donovani / Q000201:enzymology / Q000378:metabolism*; D008969:Molecular Sequence Data; D012995:Solubility; D013379:Substrate Specificity; D017855:Transferases (Other Substituted Phosphate Groups) / Q000378:metabolism*
D006028:Glycosphingolipids; C008290:lipophosphonoglycan; D017855:Transferases (Other Substituted Phosphate Groups); C108678:lipophosphoglycan Gal6-mannosylphosphoryltransferase