DnaK-facilitated ribosome assembly in Escherichia coli revisited

Abstract

Assembly helpers exist for the formation of ribosomal subunits. Such a function has been suggested for the DnaK system of chaperones (DnaK, DnaJ, GrpE). Here we show that 50S and 30S ribosomal subunits from an Escherichia coli dnaK-null mutant (containing a disrupted dnaK gene) grown at 30°C are physically and functionally identical to wild-type ribosomes. Furthermore, ribosomal components derived from mutant 30S and 50S subunits are fully competent for in vitro reconstitution of active ribosomal subunits. On the other hand, the DnaK chaperone system cannot circumvent the necessary heat-dependent activation step for the in vitro reconstitution of fully active 30S ribosomal subunits. It is therefore questionable whether the requirement for DnaK observed during in vivo ribosome assembly above 37°C implicates a direct or indirect role for DnaK in this process.

Keywords

• DnaK chaperone
• in vitro reconstitution
• ribosome assembly
• assembly helpers
• rRNA, ribosomal RNA
• TP30, total ribosomal proteins of the 30S subunit
• TP50, total ribosomal proteins of the 50S subunit
• ts, thermosensitive for growth
• tetR and cmR, genetic determinants conferring resistance to tetracycline and chloramphenicol, respectively