Trigger Factor from the Psychrophilic Bacterium

16 17 In eubacteria, trigger factor (TF) is the first chaperone to interact with newly synthe-18 sised polypeptides and assist their folding as they emerge from the ribosome. We report the 19 first characterisation of a TF from a psychrophilic organism. TF from Psychrobacter frigidi-20 cola (TF PF) was cloned, produced in Escherichia coli and purified. Strikingly, cross-linking 21 and fluorescence anisotropy analyses revealed it to exist in solution as a monomer, unlike 22 the well-characterised, dimeric E. coli TF (TF EC). Moreover, TF PF did not exhibit the down-23 turn in reactivation of unfolded GAPDH that is observed with its E. coli counterpart, even 24 at high molar ratios of trigger factor:GAPDH and revealed dramatically reduced retarda-25 tion of membrane translocation by a model recombinant protein compared with the E. coli 26 chaperone. TF PF was also significantly more effective than TF EC at increasing the yield of 27 soluble and functional recombinant protein in a cell-free protein synthesis system, indicat-28 ing it is not dependent on downstream systems for its chaperoning activity. We propose that 29 TF from P. frigidicola differs from E. coli TF in its quaternary structure and chaperone 30 activity and we discuss the potential significance of these differences in its native environ-31 ment.