High hydrostatic pressure can probe the effects of functionally related ligands on the quaternary structures of the chaperonins GroEL and GroES.
We investigated the effects of high hydrostatic pressure in the range of 1--3 kilobars on tetradecameric GroEL, heptameric GroES, and the GroEL-GroES complex. Unlike GroEL monomers formed by urea dissociation, which can be reassembled back to the tetradecamer, the pressure-dissociated monomers do not reassemble readily. This indicates an alteration of their native structures, an example of conformational drift. Pressure versus time profiles and kinetics of the dissociation of both GroEL and GroES at fixed pressures were monitored by light scattering. Unlike GroEL, GroES monomers do reassociate readily. Reaction conditions were varied by adding ATP, Mg(2+), ADP, AMP-PNP, and KCl. At any individual pressure, the dissociation process is governed by both thermodynamics and kinetics. This leads to the decrease in the yield of monomers at lower pressures. In the presence of Mg(2+) and KCl, GroEL is stable up to 3 kilobars. The presence of either ATP or ADP but not AMP-PNP leads to GroEL dissociation at lower pressures. Interestingly, the GroEL-GroES complex is very stable in the range of 1--2.5 kilobars. However, the addition of ADP destabilizes the complex, which dissociates completely at 1.5 kilobars. The results are rationalized in terms of different degrees of cooperativity between individual monomers and heptameric rings in the GroEL tetradecamer. Such allosteric interactions leading to the alteration of quaternary structure of GroEL in the absence of chemical denaturants are important in understanding the mechanism of chaperonin-assisted protein folding by the GroEL-GroES system.
Panda
M
M
Department of Biochemistry, University of Texas Health Science Center, San Antonio, Texas 78229-3900, USA.
Ybarra
J
J
Horowitz
P M
PM
D000255:Adenosine Triphosphate / Q000378:metabolism; D018835:Chaperonin 10 / Q000737:chemistry*; D018834:Chaperonin 60 / Q000737:chemistry*; D006874:Hydrostatic Pressure; D008274:Magnesium / Q000494:pharmacology; D020836:Protein Structure, Quaternary
D018835:Chaperonin 10; D018834:Chaperonin 60; D000255:Adenosine Triphosphate; D008274:Magnesium