Probing the U-Shaped Conformation of Caveolin-1 in a Bilayer

Caveolin induces membrane curvature and drives the formation of caveolae that participate in many crucial cell functions such as endocytosis. The central portion of caveolin-1 contains two helices (H1 and H2) connected by a three-residue break with both N- and C-termini exposed to the cytoplasm. Although a U-shaped configuration is assumed based on its inaccessibility by extracellular matrix probes, caveolin structure in a bilayer remains elusive. This work aims to characterize the structure and dynamics of caveolin-1 (D82-S136; Cav1(82-136)) in a DMPC bilayer using NMR, fluorescence emission measurements, and molecular dynamics simulations. The secondary structure of Cav1(82-136) from NMR chemical shift indexing analysis serves as a guideline for generating initial structural models. Fifty independent molecular dynamics simulations (100 ns each) are performed to identify its favorable conformation and orientation in the bilayer. A representative configuration was chosen from these multiple simulations and simulated for 1 mu s to further explore its stability and dynamics. The results of these simulations mirror those from the tryptophan fluorescence measurements. (i.e., Cav1(82-136) insertion depth in the bilayer), corroborate that Cav1(82-136) inserts in the membrane with U-shaped conformations, and show that the angle between H1 and H2 ranges from 35 to 69 degrees, and the tilt angle of Cav1(82-136) is 27 +/- 6 degrees. The simulations also reveal that specific faces of H1 and H2 prefer to interact with each other and with lipid molecules, and these interactions stabilize the U-shaped conformation.