In summary we described the synthesis of a series of chemically pure GDGT-0 lipids and their self-assembly into vesicles in aqueous media. We addressed the question of what conformations those lipids adopt in a membrane and how it impacts the functional behavior of the membranes. While it is generally considered that archaeal bipolar lipids adopt a membrane-spanning conformation (O-shape), we have provided structural and functional evidence that GDGT-0 lipids possess the conformational flexibility to fold into U-shapes in a membrane. First, we showed that the GDGT-0 lipids adopt U-shapes in monomolecular films at the air-water interface as evident from the significantly higher mean molecular area occupied by these lipids in comparison to monopolar lipids. Next, we showed that vesicles primarily composed of GDGT lipids can undergo pHinduced fusion with influenza virus particles at kinetic rates similar to that measured with conventional monopolar lipid vesicles. Such behavior can be explained by considering that the GDGT membranes at least partially consist of bilayer regions composed of U-shaped lipids that act as sites of fusion. Conformational flexibility is further evident from the rich mesophase behavior for each GDGT lipid which were found to transform between lamellar and non-lamellar phases at various combinations of buffer and temperature conditions. One matter that will need careful consideration in the future is the impact of the cycloalkane rings in the hydrocarbon region of the higher order GDGT-n (n ≥ 1) lipids. GDGTs containing n cyclopentane rings are expected to be conformationally rigid and less likely to loop into a U-shape. Ongoing work is focused on synthesizing such lipids to probe their structural and dynamical behavior. Supporting Information. Supplementary schemes S1-S7, Synthetic procedures, Supplementary figures S1-S12, and NMR spectral data * Steven G. Boxer: Department of Chemistry, Stanford University, Stanford CA-94305; orcid.org/0000-0001-9167-4286; Email: sboxer@stanford.edu