Substrate protein switches GroE chaperonins from asymmetric to symmetric cycling by catalyzing nucleotide exchange

The complex kinetics of Pi and ADP release by the chaperonin GroEL/GroES is influenced by the presence of unfolded substrate protein (SP). Without SP, the kinetics of Pi release are described by four phases: a lag, a burst of ATP hydrolysis by the nascent cis ring, a delay caused by ADP release from the nascent trans ring, and steady-state ATP hydrolysis. The release of Pi precedes the release of ADP. The rate-determining step of the asymmetric cycle is the release of ADP from the trans ring of the GroEL-GroES(1) bullet complex that is, consequently, the predominant species. In the asymmetric cycle, the two rings of GroEL function alternately, 180 out of phase. In the presence of SP, a change in the kinetic mechanism occurs. With SP present, the kinetics of ADP release are also described by four phases: a lag, a surge of ADP release attributable to SP-induced ADP/ATP exchange, and a pause during which symmetrical football particles are formed, followed by steady-state ATP hydrolysis. SP catalyzes ADP/ATP exchange on the trans ring. Now ADP release precedes the release of Pi, and the rate-determining step of the symmetric cycle becomes the hydrolysis of ATP by the symmetric GroEL-GroES(2) football complex that is, consequently, the predominant species. A FRET-based analysis confirms that asymmetric GroEL-GroES(1) bullets predominate in the absence of SP, whereas symmetric GroEL-GroES(2) footballs predominate in the presence of SP. This evidence suggests that symmetrical football particles are the folding functional form of the chaperonin machine in vivo.