![]() 3b), so that AFM may be well suited for our study to capture E-cad-Fc structure in cell culture. The short axis diameter of the EC domain is expected to be around 2 nm (calculated from PDB ID: 3Q2V 28, Fig. For example, the periodic double-helix structure of B-DNA in the solution was observed at a spatial resolution of 1.2 nm with the PeakForce Tapping mode 27. With AFM observation, a resolution of nm level is expected. Furthermore, there is no need for sample staining. Unlike an electron microscope, samples do not need to be dried or frozen, and can be measured in a physiological environment. AFM allows direct observation of a sample as it is. With these backgrounds, we focus on morphological observations of E-cad-Fc by using AFM (Atomic Force Microscope). It is of great importance to clarify structures of the chimeric proteins in the presence and absence of calcium ions, as well as their cell adhesion mechanism, so that the chimeric proteins may be better used for biomaterial development. Cells cultivated on plates coated with these cadherin chimeric proteins adhere in the presence of Ca 2+ and detach when Ca 2+ is removed by a chelating agent such as EDTA and EGTA. Experiments in mice aimed at developing treatments for newborn brain injury showed that N-cad-Fc-coated gelatin sponge artificial scaffolds promote neuronal regeneration and functional recovery, highlighting the significance of biomaterials also in vivo 26. Chimeric proteins of the same kind, namely N-cad-Fc and VE-cad-Fc, have been also developed, which also promote differentiation of neural progenitor cells 21, 22, 23, 24 and human umbilical vein endothelial cells (HUVECs) 25 respectively in vitro. Application of E-cad-Fc to particle coating is also in progress 20. On plates coated with E-cad-Fc, ES/iPS cells are cultivated particularly well and induced to differentiate with several methods 16, 17, 18, 19. 11, 12, 13, 14, 15 so that the chimeric proteins may serve as important biomaterial which helps clarify the cell function. E-cadherin involves in cell morphogenesis, embryonic development, cancer, EMT, etc. ![]() The EC1 and EC2 domains are the ones directly involved in the cell adhesion, and there have been many studies on the interactions between them.Į-cadherin-Fc (E-cad-Fc) is a fusion protein, which consists of the extracellular domain of E-cadherin and the IgG Fc domain 7, 8, 9, 10 (Fig. Cadherins bind by binding calcium ions to calcium ion binding sites between individual extracellular domains. 1a), and the size of one domain is about 110 amino acid residues, which are named Extra Cellular 1 (EC1) -EC5 in order from the N-terminal side. The extracellular site is composed of five domains bound in tandem (Fig. Many physiological studies have been carried out to show that cell–cell adhesion is controlled from the cytoplasmic side 2, 3, 4, 5, 6. On the cytoplasmic side, cadherins link to actin cytoskeleton through such molecules as catenin groups and EPLIN. Classical cadherins are single-pass transmembrane proteins of the cell membrane. There are more than 100 types of cadherins, among which classical cadherins are the most studied. ![]() Cadherin forms the former type of homophilic adhesion. There are two types of cell adhesion, cell–cell adhesion and cell-substrate adhesion. The biomatrix surface plays an important role in cell culture, so the analysis of its structure and function may help promote cell engineering based on cell recognition.Ĭadherin is a group of cell surface glycoproteins, identified and named as a calcium-dependent cell adhesion molecule 1. Furthermore, we succeeded in visualizing the changes in the rod-like structure of the EC domains with and without calcium. The observed structures were in good agreement with an X-ray crystallographic model. ![]() Our AFM observations revealed a rod-like structure of the entire extracellular domain of E-cad-Fc in the presence of Ca 2+ as well as trans-binding of E-cad-Fc with adjacent molecules, which may be the first, direct confirmation of trans-dimerization of E-cadherin. For the purpose of clarifying the structures of E-cad-Fc in the presence and absence of Ca 2+, we analyzed the molecular structure of E-cad-Fc by AFM in liquid. The cells adhere to the plate via E-cad-Fc in the presence of Ca 2+ and detach by a chelating agent. On plates coated with this chimeric protein, ES/iPS cells are cultivated particularly well and induced to differentiate. The fusion protein E-cad-Fc consists of the extracellular domain of E-cadherin and the IgG Fc domain. E-cadherin is a key Ca-dependent cell adhesion molecule, which is expressed on many cell surfaces and involved in cell morphogenesis, embryonic development, EMT, etc. ![]()
0 Comments
Leave a Reply. |