Expression and species-specific glycosylation of Leishmania mexicana secreted acid phosphatase in Leishmania major

Parasitic protozoa of the genus Leishmania cycle between two main developmental stages, the promastigote in the digestive tract of the sandfly and the amastigote in the mononuclear cells of the mammalian host. Both life stages have been shown to secrete a number of highly glycosylated molecules [1] and [2]. The SAP of Leishmania mexicana, a component found in large amounts in the culture supernatant of promastigotes, is the best studied example of this group of proteins. It is a copolymer of 100 kDa (SAP1) and 200 kDa (SAP2) subunits which assemble into long filaments [3]. The backbone of the filaments is formed by a ‘string-of-pearls-like’ arrangement of globular protein subunits [4] and [5]. Projecting from this filament, two types of side arms differing in length have been demonstrated by glycerol spraying/low-angle rotary metal shadowing conferring an overall ‘bottle-brush-like’ appearance [5]. These side arms correspond to the Ser/Thr-rich repeat regions of 32 and 383 amino acids of SAP1 and SAP2, respectively, which have been shown to be the exclusive targets for glycosylation via phosphoserines, and a conserved COOH-terminal domain of 54 amino acids [3] and [6]. The composition of the phosphoserine-linked glycans has been studied in detail using monoclonal antibodies directed against phosphodi- and phosphotrisaccharide subunits and oligomannose cap structures [7] and chemical structure analysis [6]. These and other studies showed that SAP shares carbohydrate epitopes with LPG, the major glycolipid component of the promastigote surface [6], [7], [8] and [9]. Here, we show that the Ser/Thr-rich repetitive region of the secreted acid phosphatase of L. mexicana serves as a target for species-specific phosphoglycosylation in Leishmania major.