CHAPERONINS GROEL AND GROES - VIEWS FROM ATOMIC-FORCE MICROSCOPY

The Escherichia coli chaperonins, GroEL and GroES, as well as their co mplexes in the presence of a nonhydrolyzable nucleotide AMP-PNP, have been imaged with the atomic force microscope (AFM). We demonstrate tha t both GroEL and GroES that have been adsorbed to a mica surface can b e resolved directly by the AFM in aqueous solution at room temperature . However, with glutaraldehyde fixation of already adsorbed molecules, the resolution of both GroEL and GroES was further improved, as all s even subunits were well resolved without any image processing. We also found that chemical fixation was necessary for the contact mode AFM t o image GroEL/ES complexes, and in the AFM images, GroEL with GroES bo und can be clearly distinguished from those without. The GroEL/ES comp lex was about 5 nm higher than GroEL alone, indicating a 2 nm upward m ovement of the apical domains of GroEL. Using a slightly larger probe force, unfixed GroEL could be dissected: the upper heptamer was remove d to expose the contact surface of the two heptamers. These results cl early demonstrate the usefulness of cross-linking agents for the deter mination of molecular structures with the AFM. They also pave the way for using the AFM to study the structural basis for the function of Gr oE system and other molecular chaperones.