Initiation factor 2 mutants promoting fast joining of ribosomal subunits in the absence of initiator tRNA or GTP

• We have previously identified several mutations in initiation factor 2 (IF2) located outside the tRNA binding domain IV of IF2 that compensate for lack of initiator tRNA (Met-tRNAi) formylation in vivo. We have also shown that these IF2 mutants promote fast joining of ribosomal subunits even when non-formylated Met-tRNAi or deacylated tRNAi was present in the 30S pre-initiation complex (30S PIC) instead of formylated fMet-tRNAi. We demonstrate here in vitro that these IF2 mutants do not require any tRNA present on the 30S subunit to promote fast subunit joining provided that GTP is present. Moreover, in the presence of fMet-tRNAi the mutants promote fast subunit joining in the presence of only GDP. Thus, A-type IF2 mutants require either GTP or fMet-tRNAi for fast subunit joining. In contrast, fast subunit joining with wild type IF2 requires the presence of both GTP and fMet-tRNAi in the 30S PIC. These results imply that the presence of tRNA on the 30S subunit is per se not required for fast subunit joining but rather for switching the 30S:IF2 complex containing wild type IF2 into its 50S docking conformation. We show also that the rate of subunit joining with A-type IF2 is much less sensitive to the energy level in the reaction mixture than the rate of subunit joining with wild type IF2. We speculate that this insensitivity of initiation to energy levels may result in deteriorated adaptation of formylation-proficient strains harbouring A-type mutations in IF2 to growth under energy-limited conditions.

Ämnesord

MEDICIN OCH HÄLSOVETENSKAP  -- Medicinsk bioteknologi -- Medicinsk bioteknologi (hsv//swe)

MEDICAL AND HEALTH SCIENCES  -- Medical Biotechnology -- Medical Biotechnology (hsv//eng)

MEDICIN OCH HÄLSOVETENSKAP  -- Medicinska och farmaceutiska grundvetenskaper -- Mikrobiologi inom det medicinska området (hsv//swe)

MEDICAL AND HEALTH SCIENCES  -- Basic Medicine -- Microbiology in the medical area (hsv//eng)

Nyckelord

protein synthesis

initiation factors

initiator tRNA

ribosome

formylation

Biochemistry

Biokemi

Bacteriology

Bakteriologi

Biokemi

Biochemistry

Mikrobiologi

Microbiology

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