CHANGING THE NATURE OF THE INITIAL CHAPERONIN CAPTURE COMPLEX INFLUENCES THE SUBSTRATE FOLDING EFFICIENCY

For the chaperonin substrates, rhodanese, malate dehydrogenase (MDH), and glutamine synthetase (GS), the folding efficiencies, and the lifet imes of folding intermediates were measured with either the nucleotide -free GroEL or the activated ATP.GroEL.GroES chaperonin complex. With both nucleotide-free and activated complex, the folding efficiency of rhodanese and MDH remained high over a large range of GroEL to substra te concentration ratios (up to 1:1). In contrast, the folding efficien cy of GS began to decline at ratios lower than 8:1. At ratios where th e refolding yields were initially the same, only a relatively small in crease (1.6-fold) in misfolding kinetics of MDR was observed with eith er the nucleotide-free or activated chaperonin complex. For rhodanese, no change was detected with either chaperonin complex. In contrast, G S lost its ability to interact with the chaperonin system at an accele rated rate (8-fold increase) when the activated complex instead of the nucleotide-free complex was used to rescue the protein from misfoldin g, Our data demonstrate that the differences in the refolding yields a re related to the intrinsic folding kinetics of the protein substrates . We suggest that the early kinetic events at the substrate level ulti mately govern successful chaperonin-substrate interactions and play a crucial role in dictating polypeptide flux through the chaperonin syst em. Our results also indicate that an accurate assessment of the trans ient properties of folding intermediates that dictate the initial chap eronin-substrate interactions requires the use of the activated comple x as the interacting chaperonin species.