DafA cycles between the DnaK chaperone system and translational machinery

DafA is encoded by the dnaK operon of Thermus thermophilus and mediates the formation of a highly stable complex between the chaperone DnaK and its co-chaperone DnaJ under normal growth conditions. DafA_Tth contains 87 amino acid residues and is the only member of the DnaK_Tth chaperone system for which no corresponding protein has yet been identified in other organisms and whose particular function has remained elusive. Here, we show directly that the DnaK_Tth-DnaJ_Tth-DafA_Tth complex cannot represent the active chaperone species since DafA_Tth inhibits renaturation of firefly luciferase by suppressing substrate association. Since DafA_Tth must be released before the substrate proteins can bind we hypothesized that free DafA_Tth might have regulatory functions connected to the heat shock response. Here, we present evidence that supports this hypothesis. We identified the 70S ribosome as binding target of free DafA_Tth. Our results show that the association of DafA_Tth and 70S ribosomes does not require the participation of DnaK_Tth or DnaJ_Tth. On the contrary, the assembly of DnaK_Tth-DnaJ_Tth-DafA_Tth and ribosome-DafA _Tth complexes seems to be competitive. These findings strongly suggest the involvement of DafA_Tth in regulatory processes occurring at a translational level, which could represent a new mechanism of heat shock response as an adaptation to elevated temperature.