Abstract
Previous studies showed that the gene for RNA polymerase subunit alpha at 72 min on the Escherichia coli chromosome is co-transcribed with genes for ribosomal proteins (r-proteins) S11, S4, and L17, and probably S13. DNA sequence analysis of a deletion mutant has now established that the S13 gene is a part of the alpha operon and the gene order is promoter (P alpha), rpsM (S13), rpsK (S11), rpsD(S4), rpoA(alpha), and rplQ(L17). The DNA sequence extending 650 bases before S13 gene was determined. In vitro transcription experiments establish the probable location of the alpha promoter (P alpha) within this sequence. The start site is 94 nucleotides upstream from the initiation codon (GUG) of the S13 gene. This promoter has features previously noted as common to E. coli promoters. However, comparison with four other sequenced promoters of r-protein operons shows no unique common features that might account for the common regulation of synthesis of r-proteins. This lack of sequence similarity in promoters of r-protein operons may be because r-protein synthesis is regulated at least partially at a post-transcriptional level.
Highlights
To look for featuresthatmight be important for coordinate control of r-protein synthesis, and differential synthesis of rproteins and RNA polymerase subunits, we have studied the DNA sequence of the promoterregion of the other transcription unitcontaining r-protein and RNApolymerase genes, the a operon
XspcZA9"Analysis of a deletion mutant, hspc2A9 (Fig. 3), has shown that thegenes for S11 and S4 are under controlof the same promoter as those for L17 and RNA polymerase subunit a [16]. (Xspc2 is another transducing phage carrying some of the same bacterial DNA as Xfus3; see Figs. 1 and 3, and Ref. 40.) It has been presumed thatthe S13 gene is part
The Right End of the 5% Eco R I Fragment-Since there are no known functions for the DNA between the end of the spc operon and the beginning of the a operon, it seemed likely that the promoter and the S13 gene would be close to the right end of the fragment
Summary
- 601 700 AGACU;GCTmCAGCATGk9CGTACA~A~AAATAGTAGGA6TGCAAT~AaGA TrGg I~tCeCACIGaTC~tTyAIGtCeAAGsGn CI lAellP~r~o~CA sCptGEAi TeCL ~AeTRAiAeGA~tThG~C~C~GIT~~t. DNA Sequencing-The sequencing method of Maxam and Gilbert polynucleotide kinase [31]. After digestion with PSt I the [29] was used, with 0.5-mm thick sequencing gels [30]. In the experi- DNA wasprecipitated with ethanol and dissolved in 50 mM Tris-HC1 ment with a labeled Pst I end, the DNAwas treated with snake (pH 8.0), 6 mM MgClZ.The DNA was digested with 5 p g / d of snake venom phosphodiesterase to increase the efficiency of labeling by venom phosphodiesterase (Sigma;0.17 unit/mg) at 37'C for 1 h. After adding EDTA to 7 a, the DNA wastreated with bacterial alkaline phosphatase and end-labeled as usual with polynucleotide kinase [29]. Run the product RNA was purified as previously described [12]
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