Zinc is a key factor in controlling alternation of two types of L31 protein in the Bacillus subtilis ribosome

Abstract

SummaryWe have analysed changes in the composition of ribosomal proteins during cell growth in Bacillus subtilis . Ribosome fractions were prepared from B. subtilis cells at different phases of growth and were separated by radical-free and highly reducing (RFHR) twodimensional polyacrylamide gel electrophoresis. We identified 50 ribosomal proteins, including two paralogues of L31 protein (RpmE and YtiA). Although the ribosome fraction extracted from exponentially growing cells contained RpmE protein, this protein disappeared during the stationary phase. In contrast, YtiA was detected in the ribosome fraction extracted after the end of exponential growth. Expression of the ytiA gene encoding YtiA was found to be negatively controlled by Zur, a zinc-specific transcriptional repressor that controls zinc transport operons. Analysis by inductively coupled plasma mass spectrometry (ICP-MS) indicated that RpmE contains one zinc ion per molecule of protein. In addition, mutagenesis of the rpmE gene encoding RpmE revealed that Cys-36 and Cys-39, located within a CxxC motif, are required not only for binding zinc but also for the accumulation of RpmE in the cell. Taken together, these results indicate that zinc plays an essential role in the alternation between two types of L31 protein in the ribosome of B. subtilis .

Towards an Elucidation of the Roles of the Ribosome during Different Growth Phases inBacillus subtilis

“…1). From these findings, together with the observation that YtiA binds to the 50S subunit of the ribosome, 62) we concluded that B. subtilis expresses two types of ribosomal L31 protein differentially and switches between them during different phases of growth.…”

“…By matrix assisted laser desorption ionization time-of-flight mass spectrometry (MALDI TOF-MS) and N-terminal sequence analyses, 51 spots were assigned to ribosomal proteins with duplicated spots for L29 (L29 and L29m, the latter detected only in the ribosomal fractions of exponentially growing cells). 62) Moreover, YvyD and YugI, which are known as general stress proteins, 63,64) were identified in the ribosomal fractions. 62) YvyD was found in the ribosomal fractions prepared from cells in stationary phase.…”

“…62) Moreover, YvyD and YugI, which are known as general stress proteins, 63,64) were identified in the ribosomal fractions. 62) YvyD was found in the ribosomal fractions prepared from cells in stationary phase. 62) Although the detailed function of YvyD was unclear when we first identified it, we have been able to elucidate the functional role of YvyD, and will report this elsewhere in the near future.…”

“…62) Ribosomal fractions extracted from exponentially-growing cells contained RpmE, but this protein disappeared during the stationary growth phase. In contrast, YtiA was detected in ribosomal fractions extracted after the end of exponential growth.…”

Graded expression of zinc-responsive genes through two regulatory zinc-binding sites in Zur

“…In many bacteria, zinc depletion causes inactivation of zinc-specific Zur to derepress zinc transporter genes (6). In Gram-positive bacteria, genes for zincless ribosomal proteins are derepressed to replace zinc-bound paralogs as a way of mobilizing zinc (7)(8)(9)(10)(11)(12). Coelibactin, a putative zincophore, has been recently reported to be regulated by Zur in Streptomyces coelicolor (13,14).…”