Monitoring protein conformation along the pathway of chaperonin-assisted folding.
The GroEL/GroES chaperonin system mediates protein folding in the bacterial cytosol. Newly synthesized proteins reach GroEL via transfer from upstream chaperones such as DnaK/DnaJ (Hsp70). Here we employed single molecule and ensemble FRET to monitor the conformational transitions of a model substrate as it proceeds along this chaperone pathway. We find that DnaK/DnaJ stabilizes the protein in collapsed states that fold exceedingly slowly. Transfer to GroEL results in unfolding, with a fraction of molecules reaching locally highly expanded conformations. ATP-induced domain movements in GroEL cause transient further unfolding and rapid mobilization of protein segments with moderate hydrophobicity, allowing partial compaction on the GroEL surface. The more hydrophobic regions are released upon subsequent protein encapsulation in the central GroEL cavity by GroES, completing compaction and allowing rapid folding. Segmental chain release and compaction may be important in avoiding misfolding by proteins that fail to fold efficiently through spontaneous hydrophobic collapse.
Sharma
Shruti
S
Department of Cellular Biochemistry, Max Planck Institute of Biochemistry, Am Klopferspitz 18, D-82152 Martinsried, Germany.
Chakraborty
Kausik
K
Müller
Barbara K
BK
Astola
Nagore
N
Tang
Yun-Chi
YC
Lamb
Don C
DC
Hayer-Hartl
Manajit
M
Hartl
F Ulrich
FU
D001419:Bacteria / Q000737:chemistry / Q000378:metabolism*; D002352:Carrier Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D018835:Chaperonin 10 / Q000378:metabolism*; D018834:Chaperonin 60 / Q000737:chemistry / Q000378:metabolism*; D031541:Fluorescence Resonance Energy Transfer; D057927:Hydrophobic and Hydrophilic Interactions; D058619:Maltose-Binding Proteins; D018832:Molecular Chaperones; D011487:Protein Conformation; D017510:Protein Folding
D002352:Carrier Proteins; D018835:Chaperonin 10; D018834:Chaperonin 60; D058619:Maltose-Binding Proteins; D018832:Molecular Chaperones