Bipolar lipids render archaeal membranes mechanical stable and impermeable even at high temperatures and at highly acidic pH. Although bipolar lipids are found in some bacteria, those are fatty acid-derived, and usually do not constitute the major lipid species in a membrane. Glycerol dialkyl glycerol tetraethers (GDGT) are one of the primary bipolar lipids found in archaea. Thermoacidophilic archaea possess GDGTs alongside other bipolar lipids while methanogenic archaea possess GDGTs alongside monopolar lipids. Fundamental understanding of structure-function relationships in GDGTs through biophysical studies will further our understanding of the basis of the adaptability of archaea to extreme environments and could aid in the design of synthetic lipids with desired properties. Archaeal bipolar lipids have long been considered as excellent building blocks of liposomes for drug delivery or artificial membranes with exceptional long-term structural and thermal stabilities. However, the scarcity of available pure GDGT lipids poses a roadblock to comprehensive biophysical studies which typically require many milligrams of lipids. Although a few model monopolar archaeal lipids in the form of ester-or ether-linked diphytan(o)yl phospholipids are commercially available, pure GDGT lipids are difficult to access. Also, the presence of multiple stereocenters make these lipids challenging to synthesize chemically. Only a handful of examples exist where polar GDGT lipids have been synthesized via total synthetic or semisynthetic approaches. To date, no GDGT lipids can be purchased from commercial vendors in pure form. Therefore, most biophysical studies until now have been performed on crude lipid extracts or model bipolar lipids. In this work we took advantage of synthetic chemistry to generate a series of four parallel GDGT-0 lipids in multi-milligram quantities (Figure ). Among these, two lipids are symmetric and two are unsymmetric in terms of head group composition. The symmetric lipids P2 and PG2 consist of phosphate and phosphoglycerol headgroups respectively on both ends. In the unsymmetric lipids P1Glc and PG1Glc a glucose group is present on one end and phosphate and phosphoglycerol groups respectively at the other end. We studied the selfassembly behavior of these GDGT lipids in various aqueous buffers and identified conditions in which membranebound giant vesicles are formed. We characterized the structural properties of GDGT membranes using small-angle X-ray scattering (SAXS) and cryogenic electron microscopy. We used SAXS also to characterize the transformations between various mesophases under a range of conditions of ionic strengths, pH, and divalent cations over a wide temperature range. Building on the physical characterization data, we decided to revisit the question of shapes of GDGT lipids in membranes. Here we offer functional evidence that the GDGT membranes contain a mixture of two kinds of molecular conformations which allows them to undergo dynamical processes such as fusion.