title: Unfolded DapA forms aggregates when diluted into free solution, confounding comparison with folding by the GroEL/GroES chaperonin system.
abstract: A recent hydrogen-deuterium exchange study of folding of the GroEL/GroES-dependent bacterial enzyme DapA has suggested that the DapA folding pathway when free in solution may differ from the folding pathway used in the presence of the GroEL/GroES chaperonin. Here, we have investigated whether DapA aggregation might be occurring in free solution under the conditions of the exchange experiment, as this would confound interpretation of the pathway predictions. Dynamic light scattering (DLS) data, sedimentation analysis and refolding yield indicate that significant aggregation occurs upon dilution of DapA from denaturant, bringing into question the earlier conclusion that different folding pathways occur in the absence and presence of the chaperonin system.
authors.affiliation: College of Pharmacy, Department of Pharmacology and Toxicology, University of Arizona, Tucson, AZ 85721.
authors.forename: Andrew
authors.initials: A
authors.lastname: Ambrose
authors.affiliation: Department of Genetics, Yale University School of Medicine, New Haven, CT 06510.
authors.forename: Wayne
authors.initials: W
authors.lastname: Fenton
authors.affiliation: College of Pharmacy, Department of Pharmacology and Toxicology, University of Arizona, Tucson, AZ 85721.
authors.forename: Damian J
authors.initials: DJ
authors.lastname: Mason
authors.affiliation: College of Pharmacy, Department of Pharmacology and Toxicology, University of Arizona, Tucson, AZ 85721.
authors.forename: Eli
authors.initials: E
authors.lastname: Chapman
authors.affiliation: Department of Genetics, Yale University School of Medicine, New Haven, CT 06510.|Howard Hughes Medical Institute, Yale University School of Medicine, New Haven, CT 06510.
authors.forename: Arthur L
authors.initials: AL
authors.lastname: Horwich
mesh_terms: D018835:Chaperonin 10 / Q000737:chemistry*; D018834:Chaperonin 60 / Q000737:chemistry*; D029968:Escherichia coli Proteins / Q000737:chemistry*; D006836:Hydro-Lyases / Q000737:chemistry*; D066329:Protein Aggregates; D017510:Protein Folding; D012996:Solutions
chemical_list: D018835:Chaperonin 10; D018834:Chaperonin 60; D029968:Escherichia coli Proteins; D066329:Protein Aggregates; D012996:Solutions; D006836:Hydro-Lyases; C020040:4-hydroxy-tetrahydrodipicolinate synthase
keywords: Aggregation; DapA; GroEL; Light scattering; Protein folding