Ligand-induced conformational changes of GroEL are dependent on the bound substrate polypeptide.
Ligand-induced conformational changes of GroEL alone and with bound rhodanese, citrate synthase, or dihydrofolate reductase were studied by limited proteolysis. Similar digestion patterns of GroEL, with or without bound substrate polypeptide, were obtained in the absence and presence of the chaperonin ligands, K+, Mg2+, or ATP. The rates of formation and degradation of the six produced proteolytic fragments were significantly different, however. Strikingly, only with Mg2+/ATP or K+/Mg2+/ATP an additional fragment of approximately 25 kDa was generated during digestion of GroEL alone or with bound rhodanese or dihydrofolate reductase, but not with bound citrate synthase. Most of the trypsin-sensitive sites in GroEL were localized in the flexible apical domain, which contains the putative polypeptide-binding region. Our data indicate that subtle structural changes in the trypsin-sensitive regions of GroEL occur as a result of the binding of the chaperonin ligands. However, these structural changes are influenced by the GroEL substrate polypeptides.
Mendoza
J A
JA
Department of Chemistry, California State University, San Marcos, California 92096-0001, USA.
Campo
G D
GD
D000255:Adenosine Triphosphate / Q000378:metabolism; D000818:Animals; D001665:Binding Sites; D002417:Cattle; D018834:Chaperonin 60 / Q000737:chemistry* / Q000378:metabolism*; D002645:Chickens; D002950:Citrate (si)-Synthase / Q000378:metabolism; D007700:Kinetics; D008024:Ligands; D008099:Liver / Q000201:enzymology; D046911:Macromolecular Substances; D008274:Magnesium / Q000378:metabolism; D008958:Models, Molecular; D009206:Myocardium / Q000201:enzymology; D010446:Peptide Fragments / Q000737:chemistry / Q000302:isolation & purification; D011188:Potassium / Q000378:metabolism; D011487:Protein Conformation; D013552:Swine; D013762:Tetrahydrofolate Dehydrogenase / Q000378:metabolism; D013884:Thiosulfate Sulfurtransferase / Q000378:metabolism; D014357:Trypsin
D018834:Chaperonin 60; D008024:Ligands; D046911:Macromolecular Substances; D010446:Peptide Fragments; D000255:Adenosine Triphosphate; D013762:Tetrahydrofolate Dehydrogenase; D002950:Citrate (si)-Synthase; D013884:Thiosulfate Sulfurtransferase; D014357:Trypsin; D008274:Magnesium; D011188:Potassium