title: Misfolded forms of glyceraldehyde-3-phosphate dehydrogenase interact with GroEL and inhibit chaperonin-assisted folding of the wild-type enzyme.
abstract: We studied the interaction of chaperonin GroEL with different misfolded forms of tetrameric phosphorylating glyceraldehyde-3-phosphate dehydrogenase (GAPDH): (1) GAPDH from rabbit muscles with all SH-groups modified by 5,5'-dithiobis(2-nitrobenzoate); (2) O-R-type dimers of mutant GAPDH from Bacillus stearothermophilus with amino acid substitutions Y283V, D282G, and Y283V/W84F, and (3) O-P-type dimers of mutant GAPDH from B. stearothermophilus with amino acid substitutions Y46G/S48G and Y46G/R52G. It was shown that chemically modified GAPDH and the O-R-type mutant dimers bound to GroEL with 1:1 stoichiometry and dissociation constants K(d) of 0.4 and 0.9 muM, respectively. A striking feature of the resulting complexes with GroEL was their stability in the presence of Mg-ATP. Chemically modified GAPDH and the O-R-type mutant dimers inhibited GroEL-assisted refolding of urea-denatured wild-type GAPDH from B. stearothermophilus but did not affect its spontaneous reactivation. In contrast to the O-R-dimers, the O-P-type mutant dimers neither bound nor affected GroEL-assisted refolding of the wild-type GAPDH. Thus, we suggest that interaction of GroEL with certain types of misfolded proteins can result in the formation of stable complexes and the impairment of chaperonin activity.
authors.affiliation: Department of Biochemistry of Animal Cell, Belozersky Institute of Physico-Chemical Biology, Moscow State University, Moscow 119992, Russian Federation.
authors.forename: Oxana V
authors.initials: OV
authors.lastname: Polyakova
authors.forename: Olivier
authors.initials: O
authors.lastname: Roitel
authors.forename: Regina A
authors.initials: RA
authors.lastname: Asryants
authors.forename: Alexei A
authors.initials: AA
authors.lastname: Poliakov
authors.forename: Guy
authors.initials: G
authors.lastname: Branlant
authors.forename: Vladimir I
authors.initials: VI
authors.lastname: Muronetz
mesh_terms: D000818:Animals; D018834:Chaperonin 60 / Q000037:antagonists & inhibitors / Q000378:metabolism*; D019281:Dimerization; D004228:Dithionitrobenzoic Acid / Q000737:chemistry; D001411:Geobacillus stearothermophilus / Q000201:enzymology; D005987:Glyceraldehyde-3-Phosphate Dehydrogenases / Q000737:chemistry* / Q000235:genetics; D009154:Mutation; D017510:Protein Folding; D011817:Rabbits; D013439:Sulfhydryl Reagents
chemical_list: D018834:Chaperonin 60; D013439:Sulfhydryl Reagents; D004228:Dithionitrobenzoic Acid; D005987:Glyceraldehyde-3-Phosphate Dehydrogenases
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