title: Unequivocal single-molecule force spectroscopy of intrinsically disordered proteins.
abstract: Intrinsically disordered proteins (IDPs) are predicted to represent about one third of the eukaryotic proteome. The dynamic ensemble of conformations of this steadily growing class of proteins has remained hardly accessible for bulk biophysical techniques. However, single-molecule techniques provide a useful means of studying these proteins. Atomic force microscopy (AFM)-based single-molecule force spectroscopy (SMFS) is one of such techniques, which has certain peculiarities that make it an important methodology to analyze the biophysical properties of IDPs. However, several drawbacks inherent to this technique can complicate such analysis. We have developed a protein engineering strategy to overcome these drawbacks such that an unambiguous mechanical analysis of proteins, including IDPs, can be readily performed. Using this approach, we have recently characterized the rich conformational polymorphism of several IDPs. Here, we describe a simple protocol to perform the nanomechanical analysis of IDPs using this new strategy, a procedure that in principle can also be followed for the nanomechanical analysis of any protein.
authors: Instituto Cajal, CSIC and Centro de Investigación Biomédica en Red sobre Enfermedades Neurodegenerativas (CIBERNED), Madrid, Spain.
authors: Javier
authors: J
authors: Oroz
authors: Rubén
authors: R
authors: Hervás
authors: Alejandro
authors: A
authors: Valbuena
authors: Mariano
authors: M
authors: Carrión-Vázquez
mesh_terms: D003001:Cloning, Molecular; D005898:Glass / Q000737:chemistry; D006801:Humans; D018625:Microscopy, Atomic Force / Q000379:methods*; D036103:Nanotechnology / Q000379:methods*; D009571:Nitrilotriacetic Acid / Q000031:analogs & derivatives / Q000737:chemistry; D009942:Organometallic Compounds / Q000737:chemistry; D010454:Peptide Termination Factors / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification; D011506:Proteins / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification
chemical_list: D009942:Organometallic Compounds; D010454:Peptide Termination Factors; D011506:Proteins; C068068:SUP35 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; C088321:nickel nitrilotriacetic acid; D009571:Nitrilotriacetic Acid