Molecular plasticity of -catenin: New insights from single-molecule measurements and MD simulation

The multifunctional protein, -catenin, has essential roles in cell adhesion and, through the Wnt signaling pathway, in controlling cell differentiation, development, and generation of cancer. Could distinct molecular forms of -catenin underlie these two functions? Our single-molecule force spectroscopy of armadillo -catenin, with molecular dynamics (MD) simulation, suggests a model in which the cell generates various forms of -catenin, in equilibrium. We find -catenin and the transcriptional factor Tcf4 form two complexes with different affinities. Specific cellular response is achieved by the ligand binding to a particular matching preexisting conformer. Our MD simulation indicates that complexes derive from two conformers of the core region of the protein, whose preexisting molecular forms could arise from small variations in flexible regions of the -catenin main binding site. This mechanism for the generation of the various forms offers a route to tailoring future therapeutic strategies

Localizing chemical groups while imaging single native proteins by high-resolution atomic force microscopy.

Simultaneous high-resolution imaging and localization of chemical interaction sites on single native proteins is a pertinent biophysical, biochemical, and nanotechnological challenge. Such structural mapping and characterization of binding sites is of importance in understanding how proteins interact with their environment and in manipulating such interactions in a plethora of biotechnological applications. Thus far, this challenge remains to be tackled. Here, we introduce force-distance curve-based atomic force microscopy (FD-based AFM) for the high-resolution imaging of SAS-6, a protein that self-assembles into cartwheel-like structures. Using functionalized AFM tips bearing Ni(2+)-N-nitrilotriacetate groups, we locate specific interaction sites on SAS-6 at nanometer resolution and quantify the binding strength of the Ni(2+)-NTA groups to histidine residues. The FD-based AFM approach can readily be applied to image any other native protein and to locate and structurally map histidine residues. Moreover, the surface chemistry used to functionalize the AFM tip can be modified to map other chemical interaction sites