title: Structural rearrangements and the unfolding mechanism of a Trigger Factor mutant studied by multiple structural probes.
abstract: Trigger Factor (TF) is a three-domain chaperone which catalyzes nascent peptide folding and harbors peptidyl-prolyl cis-trans isomerase activity. The multi-domain structure of TF makes it an interesting and challenging candidate for studies of the structural properties and functional behavior of individual domains or combined domain constructs. Here we constructed a TF mutant, NC, combining the N- and C-domains that are responsible for TF's chaperone function, and compared structural changes and unfolding characteristics of NC with wild-type TF by monitoring fluorescence spectra, far-UV CD, chemical crosslinking, DSC and binding with hydrophobic probes (ANS or bis-ANS). The results showed that the NC construct, like intact TF, could bind to hydrophobic probes, form dimers in solution, and showed a similar 3-state guanidine-induced unfolding profile. However, the NC fragment showed reduced stability towards both guanidine unfolding and thermal denaturation, suggesting that the presence of the M-domain of TF contributes to the stability of the intact TF structure.
authors.affiliation: Institute of Biophysics, Chinese Academy of Sciences, Beijing 100101, China.
authors.forename: Dong-Jie
authors.initials: DJ
authors.lastname: Fan
authors.forename: Yan-Wei
authors.initials: YW
authors.lastname: Ding
authors.forename: Jun-Mei
authors.initials: JM
authors.lastname: Zhou