Quasi-simultaneous imaging/pulling analysis of single polyprotein molecules by atomic force microscopy.
Most of studies in protein nanomechanics have used the atomic force microscope (AFM) in its force-measuring mode on immobilized protein repeats (polyproteins) as single-molecule markers. Here, we add imaging capabilities to a standard, state-of-the-art AFM "puller" and integrate the most powerful programs of analysis available for both AFM modes. This unique instrument allows high-resolution, quasi-simultaneous imaging/force spectroscopy in aqueous solution. We demonstrate its capabilities using polyproteins of a model system (titin I27 domain). This tool should greatly facilitate the development of a much needed universal functionalization system for AFM, one that should allow better sample control and an improved efficiency of protein immobilization.
Valbuena
Alejandro
A
Instituto Cajal, CSIC and Centro de Investigación Biomédica en Red sobre Enfermedades Neurodegenerativas (CIBERNED), E-28002 Madrid, Spain.
Oroz
Javier
J
Vera
Andrés Manuel
AM
Gimeno
Alejandro
A
Gómez-Herrero
Julio
J
Carrión-Vázquez
Mariano
M
D004867:Equipment Design; D019544:Equipment Failure Analysis; D007089:Image Enhancement / Q000295:instrumentation* / Q000379:methods; D008846:Micromanipulation / Q000295:instrumentation* / Q000379:methods; D018625:Microscopy, Atomic Force / Q000295:instrumentation* / Q000379:methods; D011487:Protein Conformation; D011506:Proteins / Q000737:chemistry* / Q000648:ultrastructure*; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D012815:Signal Processing, Computer-Assisted / Q000295:instrumentation*; D013048:Specimen Handling / Q000295:instrumentation* / Q000379:methods; D013314:Stress, Mechanical
D011506:Proteins