ErbB proteins belong to the epidermal growth factor (EGF) receptor family of receptor tyrosine kinases. The family has four members (ErbB1-4, also known as HER1-4), from which ErbB1, the founding member, is also called EGF receptor. ErbB1 is a receptor for EGF and other EGF-like ligands, whereas ErbB3 and ErbB4 are stimulated by different kinds of neuregulins (NRG, also known as heregulins, HRG). ErbB2 is a co-receptor for the other three members enhancing their signaling potency. According to the accepted theory, developed mainly for ErbB1, unstimulated, monomeric receptors dimerize upon growth factor binding, whose primary driving force is the conformational change induced by the ligand in the extracellular domain. The kinase domain of dimeric receptors is activated leading to transmembrane signaling.
Our work has led to the following discoveries:
The cell membrane cannot be considered a homogenous system from the standpoint of lipids or proteins since both types of molecules constitute assemblies of varying temporal and spatial stability. The hierarchical association of membrane proteins represent such a structure, but lipid microdomains also belong to them. Lipid rafts are such supramolecular organization of lipids, which are thermodynamically unstable, small (10-100 nm) structures. Proteins, lipids, the cytoskeleton and membrane turnover all contribute to their generation. They are similar, but not identical, in many respects to the liquid-ordered (Lo) domains in model membranes. The lipid environment of the cell membrane obviously influences the biophysical and cell biological properties of transmembrane proteins through their transmembrane domain. An important property of the cell membrane is the dipole potential, which is a positive potential of magnitude 200-500 mV in the interior of the cell membrane generated by the dipoles of lipids and membrane-associated water molecules. This electric field interacts with transmembrane protein (due to the dipole moment of their transmembrane domain), with ligands and molecules binding to cell surface receptors or being transported across the membrane.
We have established that
Penetratin is a cell-penetrating peptide having a potential role in efficient and targeted delivery of drugs. It enters the cytoplasm of cells either by direct membrane crossing or by endocytosis followed by traversing the membrane of the endo-lysosomal compartment (figure on the top). Due to the charged nature of penetratin, the positive intramembrane dipole potential inhibits in membrane crossing. Decreasing the dipole potential with atorvastatin treatment resulted in a significantly enhanced concentration of penetratin in the cytosol (figure at the bottom).
The cell membrane has immense medical importance since practically all drug molecules have their target in the membrane or must cross it if they have an intracellular binding site. In addition, the properties of membrane proteins are substantially influenced by the lipid composition of the cell membrane. These have medical implications for the following reasons:
We have established that
A – Labeling of the cell membrane with anti-CD14 monoclonal antibody (green) and the nucleus with DAPI (blue). Red dots corresponds to nuclei a point in the interior of a cell. B – The cell membrane (blue) and the intracellular space (red) identified using the membrane and nuclear stains. C – Overlay of the fluorescence image on the membrane and intracellular masks
Förster resonance energy transfer (FRET) is often used in the investigation of protein clustering. In FRET, a donor fluorophore passes energy an acceptor within its vicinity of 2-10 nm. Since the efficiency of the process declines with the sixth power of the donor-acceptor distance, it can be used for measuring the clustering of fluorescently labeled proteins.
Fluorescence labeling of proteins is usually achieved by fluorescent monoclonal antibodies (see figure above) or with fluorescent protein constructs. We have achieved the following results in our method development efforts:
A complete treatment of this condition is available