ATOMIC-FORCE MICROSCOPY DETECTS CHANGES IN THE INTERACTION FORCES BETWEEN GROEL AND SUBSTRATE PROTEINS

The structure of the Escherichia coli chaperonin GroEL has been invest igated by tapping-mode atomic force microscopy (AFM) under liquid. Hig h-resolution images can be obtained, which show the up-right position of GroEL adsorbed on mica with the substrate-binding site on top. Beca use of this orientation, the interaction between GroEL and two substra te proteins, citrate synthase from Saccharomyces cerevisiae with a des tabilizing Gly-->Ala mutation and RTEM beta-lactamase from Escherichia coli with two Cys-->Ala mutations, could be studied by force spectros copy under different conditions. The results show that the interaction force decreases in the presence of ATP (but not of ATP gamma S) and t hat the force is smaller for native-like proteins than for the fully d enatured ones. It also demonstrates that the interaction energy with G roEL increases with increasing molecular weight. By measuring the inte raction force changes between the chaperonin and the two different sub strate proteins, we could specifically detect GroEL conformational cha nges upon nucleotide binding.