Lack of ribosomal protein S1 in Bacillus stearothermophilus.

Abstract

The 30S ribosomal subunit of Bacillus stearothermophilus migrated as a single band when electrophoresed on agarose-acrylamide composite gels. The addition of the ribosomal protein SI purified from Escherichia coli resulted in the appearance of an additional band migrating more slowly; "Clabeled SI of E. coli was shown to be associated only with this form. Antibody against E. coli protein SI did not crossreact with either the total 30S ribosomal proteins or the postribosomal supernatant from B. stearothermophilus. These results indicate that B. stearothermophilus lacks a protein equivalent to E. coli SI and may explain our previous finding [Eur. J. Biochem. 56, 15-22 (1975)] that E.coli SI greatly stimulated the translation by B. stearothermophilus ribosomes of f2 phage RNA.In our previous paper (1) we showed that the addition of the 30S ribosomal protein SI purified from Escherichia coli caused a marked stimulation in the in vitro synthesis by Bacillus stearothermophilus ribosomes of coat protein and replicase in response to f2 phage RNA. Without this protein, B. stearothermophilus ribosomes synthesized only A-protein (maturation protein) and a trace amount of coat protein and replicase. Protein Si is the largest of all the ribosomal proteins of E. coli (2) and has been considered to be a fractional protein, that is, a protein present less than one copy per ribosome (3, 4). Recently, this protein was shown to be indispensable for the translation of natural as well as synthetic messengers (5, 6) and for the binding of MS2 phage RNA to ribosomes (7). The 30S ribosomal subunit of E. coli was shown (7-9) to exist in at least two forms, a fast migrating form (F-3OS) and a slow migrating form (S-30S), which were separable from each other by electrophoresis on agaroseacrylamide composite gels. The protein Si converted F-30S into S-30S by binding to the S'-end of 16S RNA (9). The difference between S-30S and F-SOS was thus concluded to be only the presence in the former and absence from the latter of the protein Si. MS2 phage RNA was found (7) to bind only to the former..In this paper, we report that B. stearothermophilus SOS ribosomes exist only in F-30S form and E. coli Si converts them into S-30S form. When incubated in vitro, 4CG-labeled E. coli SI was found to be associated with S-30S form and f2 phage RNA was found to bind preferentially to this form. It thus seems that the 30S ribosomal subunit of B. stearothermophilus does not have a protein functionally equivalent to E. coli Si.MATERIALS AND METHODS Preparation of Ribosomes, f2 Phage and Ribosomal SI. Ribosomes were prepared from B. stearothermophilus strain 799 and from E. coli Ai9 and RNA was extracted from f2 phage particles as described (1). The 30S ribosomal protein Si and Si-depleted 30S subunit from E. coli Ai9 were prepared as described (5, 10). The purity of Si preparation was examined by dodecylsulfate-gel electrophoresis and found more than 95% pure. Purified protein Si and B. stearothermophilus SOS ribosomes were labeled as reported (11, 12...

Expression Profiling of Translation-associated Genes in SporulatingBacillus subtilisand Consequence of Sporulation by Gene Inactivation

“…31) Inactivation of ypfD, which encodes the S1 homologue in B. subtilis, 32) also caused a considerable decrease in sporulation at high temperature (Table 3). Sorokin et al 33) reported previously that ypfD (designated it as the gene jofD) is expressed but is not essential in B. subtilis.…”

The Bacillus subtilis chromosome region encoding homologues of the Escherichia coli mssA and rpsA gene products

“…ribosomes is the apparent lack of a counterpart to the S1 ribosomal protein of Gram-negative bacteria (Higo et al, 1973(Higo et al, , 1982Isono & Isono, 1975;Schnier & Faist, 1985;Muralikrishna & Suryanarayana, 1985 ;Roberts & Rabinowitz, 1989 ; Farwell e t al., 1992). In Gram-negatives initiation of translation is facilitated by two components, one of which is a purine-rich sequence of mRNA, called the Shine-Dalgarno (SD) sequence, appropriately spaced from an initiation codon, and the second is the ribosomal protein S1, which has been shown to bind mRNA thus facilitating recognition of the initiation point (Roberts & Rabinowitz, 1989;Farwell et al, 1992).…”

Developmental expression of three proteins from the first gene of the RNA polymerase sigma 43 operon of Bacillus subtilis

“…The AG's of the ribosome-binding sites for the dnaE and rpoD genes (the second and third genes of the operon) are -13.8 (30) (31) make this explanation unlikely. On the other hand, the absence of ribosomal protein Si in B. subtilis (9) and the recent discovery of the differences between B. subtilis and E. coli in two important ribosomal protein-containing operons, i.e., the rpoD (3,15,28,29) and rpoA operons (10,26), suggest a possible difference in the ribosome components and functions that might explain the different expression patterns of the same genes as observed in our studies:…”