Membrane-mimicking environments

It is obvious why the spectroscopist wants to investigate the structure of integral membrane proteins or enzymes, whose biological action is linked to the presence of phospholipids such as phospholipase, in a membrane-mimicking environment Why such an environment should also be used for other peptides like hormones becomes more clear when we take into account the membrane compartment theory [10-12] as postulated by R. Schwyzer. This theory states that peptides that target membrane-embedded receptors... 

The tertiary structure of retrocyclin-2 in a membrane-mimicking environment was determined recently and consists of a well-defined (3-hairpin braced by the three laddered disulphide bonds.193 Analytical ultracentrifugation and NMR diffusion experiments indicated that retrocyclin-2 self-associates to form a trimer. Self-association is an important feature of some antimicrobial peptides but the role of this self-association in the mechanism of action of the retrocyclins is yet to be determined. 

Chaloin, L., Vidal, P., Heitz, A., et al. (1997) Conformations of primary amphi-pathic carrier peptides in membrane mimicking environments. Biochemistry 36, 11,179-11,187. 

Schlegel, B., Sippl, W., Holtje, H.-D. Molecular dynamics simulations of bovine rhodopsin influence of protonation states and different membrane-mimicking environments. J. Mol. Model. 2005,12, 49-64. 

Recent studies of Ananthanarayanan (122) have shown that various peptide hormones are capable of inducing Ca2+ influxes into phosphatidylcholine vesicles. The observed affinity of gastrin and CCK as well as of their lipo-derivatives for Ca2+ in membrane mimicking environments led us to examine rates of Ca2+ influxes induced by these hormones. 

Fig.l Membrane-mimicking environments, (a) Bacteriorhodopsin spin labeled at position 36 and 46 with the software MMM. (b) Schematic drawings of bacteriorhodopsin inserted in micelle, liposome, membrane bilayer and nanodisc. The liposome is not drawn to scale... 

NMR spectroscopy in a membrane-mimicking environment has been used to study the interactions of interleukin-8 with the human chemokine receptor CXCRl and how the phosphorylation of encephalins changes their structures while increasing their solubility and ability to cross the blood-brain barrier. " ... 

Piotto, S., Trapani, A., Bianchino, E., Ibarguren, M., Lopez, D.J., Busquets, X., Concilio, S. The effect of hydroxylated fatty acid-containing phospholipids in the remodeling of lipid membranes. Biochim. Biophys. Acta (BBA)-Biomembr. 1838(6), 1509-1517 (2014) Scrima, M., Di Marino, S., Grimaldi, M., Campana, F., Vitiello, G., Piotto, S.P., D Errico, G., D Ursi, A.M. Structural features of the C8 antiviral peptide in a membrane-mimicking environment. Biochim. Biophys. Acta (BBA)-Biomembr. 1838(3), 1010-1018 (2014)... 

Scrima, M., et al. Structural features of the C8 antiviral peptide in a membrane-mimicking environment. Biochim. Biophys. (BBA)-Biomembr. 1838(3), 1010-1018 (2014)... 

Fig. 2 Schematic representation of most common membrane-mimicking environments cmnpatible with solution and/or solid-state NMR studies of integral membrane proteins. Micelle (a), bicelle (b), nanolipoprotein particles also known as nanodiscs (c), and liposomes (d). The membrane protein is depicted in gray, detergents are colored in brown and lipids in blue. The apolipoprotein also known as membrane scaffold protein is represented as a green ring. Liposomes are scaled down by a factor of 2...

An alternative approach consists in reconstitution of the membrane protein in bilayer-mimicking environments, i.e., self-assembling lipid bilayer nanodiscs (Fig. lb). Nanodiscs consist of a small portion of membrane bilayer that has been solubilized by the addition of two amphipathic proteins, the membrane scaffold proteins (MSP) derived from the apolipoprotein A-1 [8-10]. Details of the preparation can also be found at http //sligarlab.life.uiuc.edu/nanodisc/protocols.html. These proteins wrap around the hydrophobic core of the lipids, effectively creating a soluble portion of membrane. 

It is emphasized that revealing the dynamics as well as the structure (or conformation) based on several types of spin-relaxation times is undoubtedly a unique and indispensable means, only available from NMR techniques at ambient temperature of physiological significance. Usually, the structure data themselves are available also from X-ray diffraction studies in a more refined manner. Indeed, better structural data can be obtained at lower temperature by preventing the unnecessary molecular fluctuations, which are major subjects in this chapter, since structural data can be seriously deteriorated for domains where dynamics are predominant even in the 2D or 3D crystalline state or proteoliposome at ambient temperature. It should be also taken into account that the solubilization of membrane proteins in detergents is an alternative means to study structure in solution NMR. However, it is not always able faithfully to mimick the biomembrane environment, because the interface structure is not always the same between the bilayer and detergent system. This typically occurs in the case of PLC-81(1-140) described in Section 4.2.4 and other types of peptide systems. 

The SPR setup can now be applied to the study of interactions between membrane associated proteins and their effectors and regulators in a membrane environment mimicking the situation in the living cell. 

Not just the enzyme but also whole enzyme catalytic systems can be mimicked as an example, if the influence of interaction of an enzyme with its environment is the target of the investigation, frequently either model membrane surfaces or aggregates formed with surfactant molecules such as micelles or vesicles are employed. 

Irrespective of the validity of the hypothetical membrane-bound pathway great efforts have been made in the last few years to determine preferred conformations of peptide hormones in media mimicking the more hydrophobic environments of membranes and/or receptor binding clefts. Our contributions in this field of research are confined to a pair of closely related gastrointestinal hormones, i.e. to gastrin and cholecystoklnin (CCK), which were chosen as model bioactive peptides. 

The dielectric constant is a bulk property of the environment of a system. The dielectric of a vacuum is one, which implies that no polarization of the environment exists. If a solvent, such as water or methanol, is explicitly included in the system, then a dielectric of one is used, because the orientation of the solvent molecules will polarize in the presence of the electric field. If water is the solvent, it is possible to use continuum models and, thus, reduce the amount of computation required. This does add artifacts to the simulation. These artifacts are fairly well characterized, however, and the computation time is dramatically reduced. In some case, two different dielectric constants were used to mimic the different biological environments. For example, in a membrane protein structural modeling, the dielectric constant was set to a value of five for simulating the hydrophobic transmembrane environment and the dielectric constant to 80 for mimicking the hydrophilic loop environment in the two phases of computer simulations (34). 

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