Mechanism of Formation and Functional Properties of Lipoprotein Discs Stabilized by Amphiphilic Maleic Acid-Containing Alternating Copolymers
Final Report Abstract
The application of copolymers in which maleic acid units alternate with (variable) hydrocarbon units (MACPs) led to an important methodological breakthrough in membrane protein research. Various MACPs were found to be able to solubilize membrane proteins from both artificial and natural lipid bilayers, forming disc-shaped lipid/lipoprotein particles enclosed by MACPs with uniform diameters in the range of 10-30 nm. The enzymes appear to remain functional in the MACP particles. Therefore, even those membrane proteins that are unstable in the presence of detergents become now amenable to techniques that require homogeneous water-soluble (single) particles. Optimized protocols of protein extraction by MACPs should yield native-like environments within MACP-lipid/lipoprotein particles. The present project aimed to draw a physico-chemical picture of formation of MACP- encased lipid/lipoprotein nanoparticles, to analyze their components interactions and to evaluate possible impact of the restricted lipid environment in these particles on the conformational dynamics of the encased membrane protein. Molecular dynamics simulations of styrene maleic acid (SMA) copolymer with lipid bilayers showed the formation of discoidal SMA lipid particles (SMALPs) with statistical and periodic 3:1 SMA copolymers and revealed that the mechanisms of interaction of SMA copolymers with lipid bilayers could be different depending on the composition and chain length of SMA copolymers. Compared to liposomes in SMALPs the lipid dynamics is restricted as probed by the reorientational dynamics of lipid bound nitroxides. In diisobutylene/maleic acid lipid particles (DIBMALPs) of different lipid composition the lipid dynamics was found to be independent from nanoparticle size, resembling that in liposomes. Photocycle and conformational dynamics of the reconstituted light sensor NpSRII/NpHtrII complex, which served here as a model system, are generally preserved in SMALPs and DIBMALPs. The protein was found to be less flexible in SMALPs with large conformational changes (millisecond time range) of the transmembrane region of NpSRII being affected. Optimizing our protocols allowed expanding the application of MACPs and lead to successful solubilization of membrane proteins expressed in mammalian or yeast cells. Thus, the full-length poreforming α-subunits hKCNH5 and hKCNQ1 of human neuronal and cardiac voltage-gated potassium channels, and the fusion construct of hKCNQ1 with its regulatory transmembrane KCNE1 β-subunit (hKCNE1-hKCNQ1) could be reconstituted into SMALPs as tetramers. Structural information of the yeast cell wall sensor Wsc1 could be obtained upon its detergent-free extraction from the yeast plasma membrane into a native-like membrane environment by SMA. Thus the application of SMALPs and DIBMALPs provide new prospects for in vitro structural, dynamic and functional studies of membrane proteins, keeping in mind possible restrictions of conformational dynamics in the transmembrane region of the encaged proteins.
Publications
- Conformational dynamics of sensory rhodopsin II in nanolipoprotein and styrene-maleic acid lipid particles. Photochem Photobiol.
Mosslehy W, Voskoboynikova N, Colbasevici A, Ricke A, Klose D, Klare JP, Mulkidzhanyan AY, Steinhoff HJ
(See online at https://doi.org/10.1111/php.13096) - Styrene/Maleic Acid Copolymers Form SMALPs by Pulling Lipid Patches out of the Lipid Bilayer. Langmuir
Orekhov PS, Bozdaganyan ME, Voskoboynikova N, Mulkidjanian AY, Steinhoff HJ, Shaitan KV
(See online at https://doi.org/10.1021/acs.langmuir.8b03978) - Lipid dynamics in nanoparticles formed by maleic acidcontaining copolymers: EPR spectroscopy and molecular dynamics simulations. BBA Biomembranes 1862
Colbasevici A, Voskoboynikova N, Orekhov PS, Bozdaganyan ME, Karlova MG, Sokolova OS, Klare JP, Mulkidjanian AY, Shaitan KV, Steinhoff HJ
(See online at https://doi.org/10.1016/j.bbamem.2020.183207) - A Three-Dimensional Model of the Yeast Transmembrane Sensor Wsc1 Obtained by SMA- Based Detergent-Free Purification and Transmission Electron Microscopy. J.Fungi 2021,7, 118
Voskoboynikova N, Karlova M, Kurre R, Mulkidjanian A.Y, Shaitan K.V, Sokolova O.S, Steinhoff HJ, Heinisch JJ
(See online at https://doi.org/10.3390/jof7020118) - Detergent-free solubilization of human Kv channels expressed in mammalian cells. Chem Phys Lipids. 219:50-57
Karlova MG, Voskoboynikova N, Gluhov GS, Abramochkin D, Malak OA, Mulkidzhanyan A, Loussouarn G, Steinhoff HJ, Shaitan KV, Sokolova OS
(See online at https://doi.org/10.1016/j.chemphyslip.2019.01.013) - Evaluation of DIBMA nanoparticles of variable size and anionic lipid content as tools for the structural and functional study of membrane proteins. BBA - Biomembranes 2021
Voskoboynikova N, Margheritis EG, Kodde F, Rademacher M, Schowe M, Budke-Gieseking A, Psathaki OE, Steinhoff HJ, Cosentino K
(See online at https://doi.org/10.1016/j.bbamem.2021.183588) - Lipid Dynamics in Diisobutylene-Maleic Acid (DIBMA) Lipid Particles in Presence of Sensory Rhodopsin II. International Journal of Molecular Sciences 2021
Voskoboynikova N, Orekhov P, Bozdaganyan M, Kodde F, Rademacher M, Schowe M, Budke- Gieseking A, Brickwedde B, Psathaki OE, Mulkidjanian AY, Cosentino K, Shaitan KV, Steinhoff HJ
(See online at https://doi.org/10.3390/ijms22052548) - Mechanisms of Formation, Structure, and Dynamics of Lipoprotein Discs Stabilized by Amphiphilic Copolymers: A Comprehensive Review. Nanomaterials 2022, 12, 361
Orekhov PS, Bozdaganyan ME, Voskoboynikova N, Mulkidjanian AY, Karlova MG, Yudenko A, Remeeva A, Ryzhykau YL, Gushchin I, Gordeliy VI, Sokolova OS, Steinhoff H-J, Kirpichnikov MP, Shaitan KV
(See online at https://doi.org/10.3390/nano12030361)
