Abstract
The gene encoding the beta subunit of Bacillus subtilis RNA polymerase was isolated from a lambda gt11 expression library using an antibody probe. Gene identity was confirmed by the similarity of its predicted product to the Escherichia coli beta subunit and by mapping an alteration conferring rifampicin resistance within the conserved rif coding region. Including the rif region, four colinear blocks of sequence similarity were shared between the B. subtilis and E. coli beta subunits. In E. coli, these conserved blocks are separated by three regions that either were not conserved or were entirely absent from the B. subtilis protein. The B. subtilis beta gene was part of a cluster with the order rplL (encoding ribosomal protein L7/L12), orf23 (encoding a 22,513-dalton protein that is apparently essential for growth), rpoB (beta), and rpoC (beta'). This organization differs from the corresponding region in E. coli by the inclusion of orf23. Experiments using promoter probe vectors and site-directed mutagenesis located a major rpoB promoter overlapping the 3'-coding region of orf23, 250 nucleotides upstream from the beta initiation codon. Thus, the B. subtilis rpoB region differs from its E. coli counterpart in both genetic and transcriptional organization.
Highlights
RNA polymerase was isolated from a Agtll expression library using an antibody probe
We identified the genes within this region by aligning the predicted B. subtilis gene products with their E. coli counterparts, using the FASTA program of Pearson and Lipman [32]
We have further shown that B. subtilis rpoB is transcribed from a promoter that overlaps the 3'-end of the preceding orf23 coding region and that this promoter is sufficiently active to support wild type growth rate and sporulation frequency
Summary
RNA polymerase was isolated from a Agtll expression library using an antibody probe. Gene identity was eonfirmed by the similarity of its predicted product to the Escherichia coli /3 subunit and by mapping an alteration conferring rifampicin resistance within the conserved rif coding region. The B. subtilis /3 gene was part of a cluster with the order rplL (encoding ribosomal protein L71L12), orf (encoding a 22,513-dalton protein that is apparently essential for growth), rpoB (/3), and rpoC (/3'). This organization differs from the corresponding region in E. coli by the inclusion of orf. The B. subtilie rpoB region differs from its E. coli counterpart in both genetic and transcriptional organization. A, the third largest subunit of the eubacterial core enzyme, has similarity to the third subunit of RNA polymerase II and to the fourth subunits of RNA polymerases I and III, this similarity is less striking than that found among the {3 or {3' homologues
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