title: Single-molecule observation of protein folding in symmetric GroEL-(GroES)2 complexes.
abstract: The chaperonin, GroEL, is an essential molecular chaperone that mediates protein folding together with its cofactor, GroES, in Escherichia coli. It is widely believed that the two rings of GroEL alternate between the folding active state coupled to GroES binding during the reaction cycle. In other words, an asymmetric GroEL-GroES complex (the bullet-shaped complex) is formed throughout the cycle, whereas a symmetric GroEL-(GroES)(2) complex (the football-shaped complex) is not formed. We have recently shown that the football-shaped complex coexists with the bullet-shaped complex during the reaction cycle. However, how protein folding proceeds in the football-shaped complex remains poorly understood. Here, we used GFP as a substrate to visualize protein folding in the football-shaped complex by single-molecule fluorescence techniques. We directly showed that GFP folding occurs in both rings of the football-shaped complex. Remarkably, the folding was a sequential two-step reaction, and the kinetics were in excellent agreement with those in the bullet-shaped complex. These results demonstrate that the same reactions take place independently in both rings of the football-shaped complex to facilitate protein folding.
authors.affiliation: Graduate School of Pharmaceutical Sciences, The University of Tokyo, 7-3-1, Hongo, Bunkyo-ku, Tokyo 113-0033, Japan.
authors.forename: Yodai
authors.initials: Y
authors.lastname: Takei
authors.forename: Ryo
authors.initials: R
authors.lastname: Iizuka
authors.forename: Taro
authors.initials: T
authors.lastname: Ueno
authors.forename: Takashi
authors.initials: T
authors.lastname: Funatsu
mesh_terms: D000251:Adenosine Triphosphatases / Q000737:chemistry; D001703:Biophysics / Q000379:methods; D018834:Chaperonin 60 / Q000378:metabolism*; D018833:Chaperonins / Q000737:chemistry; D004926:Escherichia coli / Q000378:metabolism*; D029968:Escherichia coli Proteins / Q000737:chemistry / Q000378:metabolism; D049452:Green Fluorescent Proteins / Q000737:chemistry; D007700:Kinetics; D008856:Microscopy, Fluorescence / Q000379:methods; D015233:Models, Statistical; D011485:Protein Binding; D017510:Protein Folding
chemical_list: D018834:Chaperonin 60; D029968:Escherichia coli Proteins; D049452:Green Fluorescent Proteins; D000251:Adenosine Triphosphatases; D018833:Chaperonins