PMID:15694372

Citation	Schiener, J, Witt, S, Hayer-Hartl, M and Guckenberger, R (2005) How to orient the functional GroEL-SR1 mutant for atomic force microscopy investigations. Biochem. Biophys. Res. Commun. 328:477-83
Abstract	We present high-resolution atomic force microscopy (AFM) imaging of the single-ring mutant of the chaperonin GroEL (SR-EL) from Escherichia coli in buffer solution. The native GroEL is generally unsuitable for AFM scanning as it is easily being bisected by forces exerted by the AFM tip. The single-ring mutant of GroEL with its simplified composition, but unaltered capability of binding substrates and the co-chaperone GroES, is a more suited system for AFM studies. We worked out a scheme to systematically investigate both the apical and the equatorial faces of SR-EL, as it binds in a preferred orientation to hydrophilic mica and hydrophobic highly ordered pyrolytic graphite. High-resolution topographical imaging and the interaction of the co-chaperone GroES were used to assign the orientations of SR-EL in comparison with the physically bisected GroEL. The usage of SR-EL facilitates single molecule studies on the folding cycle of the GroE system using AFM.
Links	PubMed Online version:10.1016/j.bbrc.2005.01.005
Keywords	Adsorption; Chaperonin 60/analysis; Chaperonin 60/chemistry; Chaperonin 60/ultrastructure; Micromanipulation/methods; Microscopy, Atomic Force/methods; Mutagenesis, Site-Directed; Protein Binding; Protein Conformation; Protein Folding; Structure-Activity Relationship
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