Translation Control of Swarming Proficiency in Bacillus subtilis by 5-Amino-pentanolylated Elongation Factor P

Abstract

Elongation factor P (EF-P) accelerates diprolyl synthesis and requires a posttranslational modification to maintain proteostasis. Two phylogenetically distinct EF-P modification pathways have been described and are encoded in the majority of Gramnegative bacteria, but neither is present in Gram-positive bacteria. Prior work suggested that the EF-P-encoding gene (efp) primarily supports Bacillus subtilis swarming differentiation, whereas EF-P in Gram-negative bacteria has a more global housekeeping role, prompting our investigation to determine whether EF-P is modified and how it impacts gene expression in motile cells. We identified a 5-aminopentanol moiety attached to Lys 32 of B. subtilis EF-P that is required for swarming motility. A fluorescent in vivo B. subtilis reporter system identified peptide motifs whose efficient synthesis was most dependent on 5-aminopentanol EF-P. Examination of the B. subtilis genome sequence showed that these EF-P-dependent peptide motifs were represented in flagellar genes. Taken together, these data show that, in B. subtilis, a previously uncharacterized posttranslational modification of EF-P can modulate the synthesis of specific diprolyl motifs present in proteins required for swarming motility.

Carbonyl reduction by YmfI in Bacillus subtilis prevents accumulation of an inhibitory EF‐P modification state

“…EF‐P proteins are thought to be activated by post‐translational modification and B. subtilis EF‐P is post‐translationally modified by a 5‐aminopentanol group attached to lysine 32 (Rajkovic et al, ). B. subtilis EF‐P resolved as two bands in semi‐native gel electrophoresis and Western blot analysis, and we hypothesized that the two bands could represent different states of EF‐P modification and/or activity (Fig.…”

“…In nearly all systems studied to date, depletion of EF‐P is either lethal or results in severe growth limitation, presumably due to the impaired ability to translate target sequences in essential genes (Schnier et al, ; Patel et al, ; Balibar et al, ; Yanagisawa et al, ). The one exception thus far is the Gram positive firmicute bacterium Bacillus subtilis where EF‐P is not required for growth and is instead specifically required for swarming motility, a flagellar‐mediated multicellular behavior in which cells rapidly move across a semi‐solid surface (Kearns et al, ; Rajkovic et al, ).…”

“…B. subtilis modifies EF‐P lysine 32 with yet another chemical variant, 5‐aminopentanol. Substitution of the conserved modified lysine to alanine (EF‐P K32A ) was shown to impair surface motility but the enzymes required for 5‐aminopentanolylation were unknown (Rajkovic et al, ). Here, we revisit the swarming mutants from the same screen that first discovered EF‐P as a swarming motility activator and we identify YmfI as a protein required for EF‐P 5‐aminopentanolylation.…”

Structural basis for EarP-mediated arginine glycosylation of translation elongation factor EF-P

“…Activation of the proline-specific translation elongation factors EF-P and IF-5A is usually achieved by post-translational elongations of the ε-amino group of a conserved lysine. [18][19][20][21][49][50] The resultant noncanonical amino acidsβ-lysinyl-hydroxylysine, hypusine and 5-amino-pentanolyl-lysine -appear to be chemically and structurally analogous. We recently showed that in a subset of bacteria a so far unappreciated form of post-translational modification plays an important role in the activation of EF-P.…”

Rewiring protein synthesis: From natural to synthetic amino acids

“…In vivo , a specialized elongation factor (EF-P in bacteria and eIF5A in eukaryotes) promotes efficient translation of Pro codons [69–71]. Enhancement of Pro translation by EF-P or eIF5A requires diverse post-translational modifications, including β-lysylation in E. coli and Salmonella [72, 73], rhamnosylation in Pseudomonas auruginosa [74], lysine modification with 5-aminopentanol group in Bacillus [75], and hypusine in yeast [71]. …”