Catalysis, commitment and encapsulation during GroE-mediated folding.
The Escherichia coli GroE chaperones assist protein folding under conditions where no spontaneous folding occurs. To achieve this, the cooperation of GroEL and GroES, the two protein components of the chaperone system, is an essential requirement. While in many cases GroE simply suppresses unspecific aggregation of non-native proteins by encapsulation, there are examples where folding is accelerated by GroE. Using maltose-binding protein (MBP) as a substrate for GroE, it had been possible to define basic requirements for catalysis of folding. Here, we have analyzed key steps in the interaction of GroE and the MBP mutant Y283D during catalyzed folding. In addition to high temperature, high ionic strength was shown to be a restrictive condition for MBP Y283D folding. In both cases, the complete GroE system (GroEL, GroES and ATP) compensates the deceleration of MBP Y283D folding. Combining kinetic folding experiments and electron microscopy of GroE particles, we demonstrate that at elevated temperatures, symmetrical GroE particles with GroES bound to both ends of the GroEL cylinder play an important role in the efficient catalysis of MBP Y283D refolding. In principle, MBP Y283D folding can be catalyzed during one encapsulation cycle. However, because the commitment to reach the native state is low after only one cycle of ATP hydrolysis, several interaction cycles are required for catalyzed folding.
Beissinger
M
M
Institut für Biophysik und Physikalische Biochemie, Universität Regensburg, Regensburg, 93040, Germany.
Rutkat
K
K
Buchner
J
J
D018528:ATP-Binding Cassette Transporters; D000251:Adenosine Triphosphatases / Q000378:metabolism; D000255:Adenosine Triphosphate / Q000378:metabolism; D001081:Apyrase; D001224:Aspartic Acid; D001426:Bacterial Proteins / Q000378:metabolism*; D002352:Carrier Proteins / Q000737:chemistry / Q000378:metabolism*; D002384:Catalysis; D018835:Chaperonin 10 / Q000378:metabolism; D018834:Chaperonin 60 / Q000378:metabolism; D018833:Chaperonins; D004926:Escherichia coli / Q000378:metabolism*; D029968:Escherichia coli Proteins; D006360:Heat-Shock Proteins / Q000378:metabolism*; D058619:Maltose-Binding Proteins; D008854:Microscopy, Electron; D009004:Monosaccharide Transport Proteins; D017510:Protein Folding; D012965:Sodium Chloride; D012996:Solutions; D014443:Tyrosine
D018528:ATP-Binding Cassette Transporters; D001426:Bacterial Proteins; D002352:Carrier Proteins; D018835:Chaperonin 10; D018834:Chaperonin 60; D029968:Escherichia coli Proteins; C028715:GroE protein, E coli; D006360:Heat-Shock Proteins; D058619:Maltose-Binding Proteins; D009004:Monosaccharide Transport Proteins; D012996:Solutions; C461599:maltose transport system, E coli; D001224:Aspartic Acid; D014443:Tyrosine; D012965:Sodium Chloride; D000255:Adenosine Triphosphate; D000251:Adenosine Triphosphatases; D018833:Chaperonins; D001081:Apyrase