title: Two distinct intermediates of trigger factor are populated during guanidine denaturation.
abstract: Trigger factor (TF) is an important catalyst of nascent peptide folding and possesses both peptidyl-prolyl cis-trans isomerase (PPIase) and chaperone activities. TF has a modular structure, containing three domains with distinct structural and functional properties. The guanidine hydrochloride (GuHCl) induced unfolding of TF was investigated by monitoring Trp fluorescence, far-UV CD, second-derivative UV absorption, enzymatic and chaperone activities, chemical crosslinking and binding of the hydrophobic dye, 1-anilinonaphthalene-8-sulfonate (ANS); and was compared to the urea induced unfolding. The native state of TF was found to bind ANS in 1:1 stoichiometry with a K(d) of 84 microM. A native-like state, N', is stable around 0.5 M GuHCl, and shows increased ANS binding, while retaining PPIase activity and most secondary and tertiary structure, but loses chaperone and dimerization activities, consistent with slight conformational rearrangement. A compact denatured state, I, is populated around 1.0 M GuHCl, is inactive and does not show significant binding to ANS. The data suggest that TF unfolds in a stepwise manner, consistent with its modular structure. The ability of TF to undergo structural rearrangement to maintain enzymatic activity while reducing chaperone and dimerization abilities may be related to the physiological function of TF.
authors.affiliation: National Laboratory of Biomacromolecules, Institute of Biophysics, Chinese Academy of Sciences, 15 Datun Road, Beijing 100101, China.
authors.forename: Chuan-Peng
authors.initials: CP
authors.lastname: Liu
authors.forename: Zhen-Yu
authors.initials: ZY
authors.lastname: Li
authors.forename: Guo-Chang
authors.initials: GC
authors.lastname: Huang
authors.forename: Sarah
authors.initials: S
authors.lastname: Perrett
authors.forename: Jun-Mei
authors.initials: JM
authors.lastname: Zhou
mesh_terms: D000817:Anilino Naphthalenesulfonates; D002942:Circular Dichroism; D003432:Cross-Linking Reagents; D019281:Dimerization; D004591:Electrophoresis, Polyacrylamide Gel; D029968:Escherichia coli Proteins / Q000737:chemistry* / Q000187:drug effects*; D019791:Guanidine / Q000494:pharmacology*; D018832:Molecular Chaperones; D009113:Muramidase / Q000737:chemistry; D019696:Peptidylprolyl Isomerase / Q000737:chemistry* / Q000187:drug effects*; D011489:Protein Denaturation; D017510:Protein Folding; D017434:Protein Structure, Tertiary / Q000187:drug effects; D013050:Spectrometry, Fluorescence; D013388:Succinimides; D013816:Thermodynamics
chemical_list: D000817:Anilino Naphthalenesulfonates; D003432:Cross-Linking Reagents; D029968:Escherichia coli Proteins; D018832:Molecular Chaperones; D013388:Succinimides; C027132:1-anilino-8-naphthalenesulfonate; D009113:Muramidase; C403324:trigger factor, E coli; D019696:Peptidylprolyl Isomerase; D019791:Guanidine; C019358:disuccinimidyl suberate