| Numerous molecular chaperones functions in diverse aspects of biochemical process inall kingdoms of life, including bacteria, yeast and mammals, which form a complex androbust network. These chaperones act in various cellular processes, including nascentpeptides folding, protein complex assembly, protein transport and secretion. Furthermore,chaperones also play crucial roles in protein control. Chaperones can disaggregate proteinaggregates and unfold the misfolded proteins, which will contribute to proteolytic degradation.The network consisting of various chaperones plays important roles in the proteostasismaintenance.Trigger Factor (TF) is an abundant~50kDa protein and an essential member inprokaryotic chaperones network, which binds to ribosomes and interacts with most nascentpolypeptides. The crystal structure of E. Coli TF reveals an elongated structure consisting of twomodules: peptidyl-prolyl-cis/trans-isomerase (PPIase) domain and a Crevice constructed by bothN-and C-terminal domains. While the Crevice is demonstrated to provide a protective spaceover the peptide exit site of ribosome for nascent polypeptides to fold, TF may remain bound tocertain polypeptides after their release from the ribosome, which is consistent with a role for TFas a holdasein ribosomeassembly.Based on multiple sequence alignment of TF sequences from different species and theanalysis of TF structure, two hydrophobic regions in PPIase domain and C-terminal domainswere chosen as the potential chaperone site. To investigate the effect of the hydrophobic regionsin chaperone function, here, we construct a series of point and truncation mutants, includingI195G, F233G, F322S, L336S/L337G, TF389and NC, and measured their chaperoneactivities, using lysozyme, BCAII and GAPDH as substrate. our results showed that themutation and truncation in the chosen regions had significant impacts on the chaperone functionof TF, which indicated that these two hydrophobic regions were involved in substrate recognitionofTF. Our studies will contribute to theunderstanding of structureand function ofTF. |
