Caveolin-1 is the most important protein found in caveolae， which are cell surface invaginations of the plasma membrane that act as signaling platforms. A single point mutation in the transmembrane domain of caveolin-1 (proline 132 to leucine) has deleterious effects on caveolae formation in vivo and has been implicated in various disease states， particularly aggressive breast cancers. Using a combination of gel filtration chromatography and analytical ultracentrifugation， we found that a fully functional construct of caveolin-1 (Cav1(62-178)) was a monomer in dodecylphosphocholine micelles. In contrast， the P132L mutant of Cav1(62-178) was dimeric. To explore the dimerization of the P132L mutant furthe... More
Caveolin-1 is the most important protein found in caveolae， which are cell surface invaginations of the plasma membrane that act as signaling platforms. A single point mutation in the transmembrane domain of caveolin-1 (proline 132 to leucine) has deleterious effects on caveolae formation in vivo and has been implicated in various disease states， particularly aggressive breast cancers. Using a combination of gel filtration chromatography and analytical ultracentrifugation， we found that a fully functional construct of caveolin-1 (Cav1(62-178)) was a monomer in dodecylphosphocholine micelles. In contrast， the P132L mutant of Cav1(62-178) was dimeric. To explore the dimerization of the P132L mutant further， various truncated constructs (Cav1(82-178)， Cav1(96-178)， Cav1(62-136)， Cav1(82-136)， Cav1(96-136)) were prepared which revealed that oligomerization occurs in the transmembrane domain (residues 96-136) of caveolin-1. To characterize the mutant structurally， solution-state NMR experiments in lyso-myristoylphosphatidylglycerol were undertaken of the Cav1(96-136) P132L mutant. Chemical shift analysis revealed that， compared to the wild-type， helix 2 in the transmembrane domain was lengthened by four residues (wild-type， residues 111-129; mutant， residues 111-133)， which corresponds to an extra turn in helix 2 of the mutant. Lastly， point mutations at position 132 of Cav1(62-178) (P132A， P132I， P132V， P132G， P132W， P132F) revealed that no other hydrophobic amino acid can preserve the monomeric state of Cav1(62-178)， which indicates that proline 132 is critical in supporting proper caveolin-1 behavior.
