A broadly active fucosyltransferase LmjFUT1 whose mitochondrial localization and catalytic activity is essential in the parasitic protozoan Leishmania

Glycoconjugates play major roles in the infectious cycle of the trypanosomatid parasite Leishmania. While GDP-Fucose synthesis is essential (Guo et al 2017), fucosylated glycoconjugates have not been reported in Leishmania major. Four predicted fucosyltransferases appear conventionally targeted to the secretory pathway; SCA1/2 play a role in side-chain modifications of lipophosphoglycan, while gene deletion studies here showed that FUT2 and SCAL were not essential. Unlike most eukaryotic glycosyltransferases, the predicted α 1-2 fucosyltransferase encoded by FUT1 localized to the mitochondrion. A quantitative plasmid segregation assay, expressing FUT1 from the multicopy episomal pXNG vector in a chromosomal null fut1- background, established that FUT1 is essential. Similarly plasmid shuffling confirmed that both enzymatic activity and mitochondrial localization were required for viability, comparing import-blocked or catalytically inactive enzymes respectively. Enzymatic assays of tagged proteins expressed in vivo or of purified recombinant FUT1 showed it had a broad fucosyltransferase activity including glycan and peptide substrates. Unexpectedly a single rare fut1 s segregant was obtained in rich media, which showed severe growth defects accompanied by mitochondrial dysfunction and loss, all of which were restored upon FUT1 re-expression. Thus, FUT1 along with the similar Trypanosoma brucei enzyme TbFUT1 (Bandini et al 2021) joins the eukaryotic O-GlcNAc transferase isoform as one of the few glycosyltransferases acting within the mitochondrion. Trypanosomatid mitochondrial FUT1s may offer a facile system for probing mitochondrial glycosylation in a simple setting and their essentiality renders it an attractive target for chemotherapy of these serious human pathogens.