BAR domain protein assemblies are proposed to be stabilized by amphipathic helices as in endophilin N-BAR domains, or through edge-edge interactions such as in various F-BAR domains. The intermolecular contacts in F-BAR domains are generated through charged residues at the contact interfaces. Since residue D151 is located in the arm region with Saponin-V the side chain pointing outward in the crystal structure, this orientation might allow for interaction with a charged residue from a neighboring BAR domain in an anti-parallel manner. The crystal structure reveals several charged or polar residues at the distal tip region. We screened these residues by mutation to alanines and transfected the resulting mutants in C2C12 cells to determine if they were important for maintenance of tubulation capacity as well. We first recall that the K35 residue is predicted to be Liriope-muscari-baily-saponins-C projected onto the charged surface of the N-terminal helix in BIN1, thus the K35N mutation may influence helix insertion into the lipid membrane and influence membrane tubulation. To test this, we compared the interaction of WT and K35N BAR domain with lipid monolayers. The lipid monolayer was spread at a constant area at a given initial surface pressure, and the change in surface pressure was monitored on a Langmuir trough after injection of proteins into the subphase. A linear relationship between Dp and p0 was observed that allows determination of the critical penetration pressure pc, which can be interpreted as the upper limit of p0 that allows protein penetration into the lipid membrane. In conclusion, our studies have shown that both protein density and oligomerization on membranes determine membrane curvature generation ability. Based on the membrane structures revealed in the tubulation assay, we suggest that in order to initiate spontaneous liposome deformation and tubule growth, transient ordered protein oligomers are required to form on a flat membrane and to allow for the initiation of tubule formation. We have experimentally shown that protein-protein assembly is required to drive membrane bud formation in the early stage of membrane deformation which is consistent with the results from simulations.