Giardia lamblia, is an intestinal protozoan parasite responsible as a major cause of water-borne intestinal infection in humans (giardiasis). It has been proposed earlier that Giardia has limited lipid synthesis ability and therefore relies on preformed lipid molecules from outside sources. It has also been proposed that lipids are taken up through passive and active transport, and giardial flippase play a significant role in this process. This study describes the identification and characterization of a putative homologue of flippase gene from Giardia. Flippase is an enzyme that functions as a trans-bilayer transport system facilitating the flip flop diffusion of phospholipids that are generated and or modified within the cell to the outer face of the lipid bilayer. Flippase can also actively transport aminophospholipids from the outer membrane to the inner monolayer Additionally, eukaryotic flippase has been implicated in the movement of synthesized membrane lipids among organelles. Beginning with the putative gene sequence from the Giardia genome, this study verifies the expression of the gene in trophozoites, expresses recombinant proteins for immunologic studies, and constructs a knockout flippase mutant to test for functional dependence of Giardia on this gene. I speculate that flippase plays a potentially vital role in phospholipid transport and provides potential for novel chemotherapeutic treatments against G. lamblia.Oligos were extended to profile unique restriction sites for cloning and ordered from MWG Biotech (Cork, Ireland) as depicted in Table- ... PCR products were cloned using a TOPO TA cloning kit from Invitrogen Corporation, (Grand Island, NY).
|Title||:||Giardia Lamblia: Genomic and Molecular Analyses of Flippase|
|Author||:||Diana L. Villazana-Kretzer|
|Publisher||:||ProQuest - 2008|