Polyols induce ATP-independent folding of GroEL-bound bacterial glutamine synthetase.
We have previously assessed the GroE chaperonin requirements for folding of bacterial glutamine synthetase (GS) and established that, at 37 degrees C in 50 mM Tris buffer, ATP binding to the GroEL-GS complex is mandatory for the release and reactivation of dodecameric enzyme. However, we demonstrate here that the addition of 1-4 M glycerol to GroEL-GS complexes resulted in release and reactivation of GS in the absence of nucleotide. Furthermore, the kinetics of refolding and refolding yields of this glycerol-induced refolding were similar to those observed with ATP. Other polyols such as sucrose, 1,2-propanediol, or 1,3-propanediol also facilitated nucleotide-independent refolding of GS from chaperonin complex. The observed phenomenon cannot be attributed to the viscosity or molecular crowding effects because solutions of dextran or Ficoll with the same viscosity as 4 M glycerol failed to reactivate GroEL-bound GS. Like glycerol, other osmolytes such as betaine and sarcosine or high salt (500 mM NaCl) facilitated spontaneous folding of GS. However, no reactivation of GroEL-bound GS was observed with these additives. The presence of glycerol affected binding of fluorescent probe 1,8-anilinonaphthalene to GroEL, suggesting that glycerol may alter the chaperonin structure. Our data suggest that low-molecular-weight polyols affect both GroEL and bound GS monomers to reduce their binding affinity. This results in an increased partitioning of GS toward active, assembly-competent states.
Voziyan
Paul A
PA
Department of Biochemistry and Molecular Biology, University of Kansas Medical Center, Kansas City, Kansas 66160-7421, USA.
Fisher
Mark T
MT
D000255:Adenosine Triphosphate / Q000378:metabolism*; D000438:Alcohols / Q000494:pharmacology*; D000817:Anilino Naphthalenesulfonates / Q000378:metabolism; D001426:Bacterial Proteins / Q000378:metabolism; D018834:Chaperonin 60 / Q000378:metabolism*; D005974:Glutamate-Ammonia Ligase / Q000378:metabolism*; D005990:Glycerol / Q000494:pharmacology; D019946:Propylene Glycol / Q000494:pharmacology; D017510:Protein Folding; D012996:Solutions; D013395:Sucrose / Q000494:pharmacology
D000438:Alcohols; D000817:Anilino Naphthalenesulfonates; D001426:Bacterial Proteins; D018834:Chaperonin 60; D012996:Solutions; D013395:Sucrose; C027132:1-anilino-8-naphthalenesulfonate; D019946:Propylene Glycol; D000255:Adenosine Triphosphate; D005974:Glutamate-Ammonia Ligase; D005990:Glycerol