Identification, biochemical characterization, and in-vivo expression of the intracellular invertase BfrA from the pathogenic parasite Leishmania major.
The parasitic life cycle of Leishmania includes an extracellular promastigote stage that occurs in the gut of the insect vector. During that period, the sucrose metabolism and more specifically the first glycosidase of this pathway are essential for growth and survival of the parasite. We investigated the expression of the invertase BfrA in the promastigote and amastigote stages of three parasite species representative of the three various clinical forms and of various geographical areas, namely Leishmania major, L. donovani and L. braziliensis. Thereafter, we cloned, overexpressed and biochemically characterized this invertase BfrA from L. major, heterologously expressed in both Escherichia coli and L. tarentolae. For all species, expression levels of BfrA mRNA were correlated to the time of the culture and the parasitic stage (promastigotes > amastigotes). BfrA exhibited no activity when expressed as a glycoprotein in L. tarentolae but proved to be an invertase when not glycosylated, yet owing low sequence homology with other invertases from the same family. Our data suggest that BfrA is an original invertase that is located inside the parasite. It is expressed in both parasitic stages, though to a higher extent in promastigotes. This work provides new insight into the parasite sucrose metabolism.
Laboratory of Parasitology-Mycology, Centre Hospitalier Universitaire de Rennes, France; INSERM U1085/IRSET - Institut de Recherche en Santé, Environnement et Travail, Université Rennes 1, France.
Sorya
S
Belaz
Univ. Orléans, CNRS, ICOA, UMR 7311, F-45067 Orléans, France.
Thibault
T
Rattier
Univ. Orléans, CNRS, ICOA, UMR 7311, F-45067 Orléans, France.
Pierre
P
Lafite
Univ. Orléans, CNRS, ICOA, UMR 7311, F-45067 Orléans, France.
Philippe
P
Moreau
Department of Cellular Chemistry, Hannover Medical School, Carl-Neuberg-Strasse 1, 30625 Hannover, Germany.
Françoise H
FH
Routier
Laboratory of Parasitology-Mycology, Centre Hospitalier Universitaire de Rennes, France; INSERM U1085/IRSET - Institut de Recherche en Santé, Environnement et Travail, Université Rennes 1, France.
Florence
F
Robert-Gangneux
Laboratory of Parasitology-Mycology, Centre Hospitalier Universitaire de Rennes, France; INSERM U1085/IRSET - Institut de Recherche en Santé, Environnement et Travail, Université Rennes 1, France.
Jean-Pierre
JP
Gangneux
Univ. Orléans, CNRS, ICOA, UMR 7311, F-45067 Orléans, France. Electronic address: richard.daniellou@univ-orleans.fr.
Richard
R
Daniellou
D000818:Animals; D004926:Escherichia coli; D007313:Insecta / Q000469:parasitology; D007892:Leishmania braziliensis / Q000378:metabolism; D007893:Leishmania donovani / Q000378:metabolism; D018320:Leishmania major / Q000201:enzymology* / Q000378:metabolism; D008958:Models, Molecular; D012333:RNA, Messenger / Q000378:metabolism*; D017422:Sequence Analysis, DNA; D013395:Sucrose / Q000378:metabolism; D043324:beta-Fructofuranosidase / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*
D012333:RNA, Messenger; D013395:Sucrose; D043324:beta-Fructofuranosidase
Glycosylation; Invertase; Leishmania; Leishmaniasis; Sucrose