Biology subjects

Glover, K. J.

Publications and source records attributed to Glover, K. J..

3 recordsLinked to original sources

bioRxiv2025

Proline 110 is necessary for maintaining a compact helical arrangement in caveolin-1

Caveolin-1 (Cav1) is an integral membrane protein essential for the formation of caveolae, plasma microdomains implicated in signal transduction and mechanoprotection. Cav1 is comprised of three major alpha helices, but the topology these helices adopt remains unclear. Proline 110 is located between helix 1 and helix 2, and is hypothesized to enable Cav1 to adopt an intramembrane turn crucial for the cytosolic topology of Cav1. To assess the structural role of Proline 110, we utilized Forster resonance energy transfer (FRET) between native tryptophan (W128) and site-specifically labeled dansyl fluorophores to monitor conformational changes induced by the mutation of Proline 110 to Alanine (P110A). Static light scattering confirmed that all FRET constructs behaved monomerically, ensuring intramolecular energy transfer measurements. Our results show a significant decrease in FRET efficiency upon the P110A mutation consistent with a large conformational change. These findings support the critical role of P110 in maintaining the native topology of Cav1 and highlights the structural sensitivity of the intramembrane turn.

biochemistry↗

bioRxiv2025

Interhelical distance measurements support a hairpin conformation for Caveolin-1 in phospholipid bicelles

Previous molecular dynamics (MD) simulations of caveolin-1 (Cav1) in our labs revealed the possibility of two stable conformations of its intramembrane helices (H1 and H2). To distinguish between these two conformations, experimental intramolecular distances obtained using FRET (Forster resonance energy transfer) were integrated into the MD simulations to better define the position of these helices. Two mutants of Cav1 were generated where an acceptor fluorophore, dansyl, was positioned at the N-terminal end and center of H1 (A87C, F99C respectively), while a donor fluorophore, a native tryptophan, was positioned at the C-terminal end of H2 (W128). For the A87C W128 mutant, a distance of 22.5 {+/-} 0.3 [A] was observed while a distance of 24.4 {+/-} 0.2 [A] was observed for the F99C W128 mutant in phospholipid bicelles. These experimental FRET distances were compared to distances in MD simulations of over 100 Cav1 structures. Together these studies support that H1 and H2 adopt a hairpin conformation in bicelles.

biochemistry↗

bioRxiv2018

A novel lipid-polymer system with unique properties has potential in drug delivery and biotechnology applications

Bicelles have been demonstrated to be a valuable tool for studying membrane protein interactions and structure in vitro. They are distinguished by a distinct lipid bilayer that mimics the plasma membrane of cells making it more native-like than its detergent micelle counter-part. Bicelles are typically comprised of a long-chain phospholipid such as dimyristoylphosphatidylcholine (DMPC) and a short-chain phospholipid such as dihexanoylphosphatidylcholine (DHPC). When mixed together in solution DMPC-DHPC bicelles assume a discoidal structure comprised of a heterogeneous arrangement where the short-chain lipids gather around the rim of the disk and the long-chain lipids form the flat, planar, bilayer region. In this study, the nonionic surfactant, C8E5, was used to prepare mixtures with DMPC to determine if it adopts properties similar to bicelles with a q [≥] 0.5. At q [≥] 0.5, DMPC-DHPC bicelles are bilayered and DMPC is sequestered from the detergent micelle-like DHPC. Mixtures of DMPC and C8E5 were prepared at various q values, a parameter used to describe the mole ratio of DMPC to DHPC in the preparation of bicelles. Employing biophysical methods like dynamic light scattering, 31P-NMR and analytical ultracentrifugation, properties of these lipid-detergent complexes are described. Interestingly they adopted a spherical-shaped micellar structure morphology and did not assume a discoidal shape typical of bicelles at q [≥] 0.5. However, they appear to retain bilayer-like properties that may prove beneficial for in vitro biophysical studies of membrane proteins.

biophysics↗