We have been characterizing RNA polymerase holoenzymes from Rhodobacter sphaeroides. RNA polymerase purified from R. sphaeroides transcribed from promoters recognized by Escherichia coli E sigma 32 or E sigma 70 holoenzyme. Antisera to E. coli sigma 32 or sigma 70 indicated that related polypeptides of approximately 37 kDa (sigma 37) and 93 kDa (sigma 93), respectively, are present in this preparation. Transcription of sigma 32-dependent promoters was observed in a further fractionated R. sphaeroides holoenzyme containing the sigma 37 polypeptide, while a preparation enriched in sigma 93 transcribed sigma 70-dependent promoters. To demonstrate further that the sigma 93 polypeptide functions like E. coli sigma 70, we obtained an R. sphaeroides E sigma 93 holoenzyme capable of transcription from sigma 70-dependent promoters by combining sigma 93 with (i) an E sigma 37 fraction with diminished sigma 93 polypeptide content or (ii) E. coli core RNA polymerase. The generation of analogous DNase I footprints on the lacUV5 promoter by R. sphaeroides E sigma 93 and by E. coli E sigma 70 suggests that the overall structures of these two holoenzymes are similar. However, some differences in promoter specificity between R. sphaeroides E sigma 93 and E. coli E sigma 70 exist because transcription of an R. sphaeroides rRNA promoter was detected only with E sigma 93.