FIG. 5.

Transcription of the pdhABCD operon. The genes in the pdhABCD operon encode the pyruvate decarboxylase E1α and E1β, dihydrolipoamide acetyltransferase E2, and dihydrolipoamide dehydrogenase E3 subunits of the pyruvate dehydrogenase multienzyme complex (PDH), respectively. Transcription of this operon results in a 5.2-kb pdhABCD mRNA (13). The 5′ end of the pdhABCD transcript was determined by primer extension analysis (lower panel). Total RNAs from cells of strain 1A765 (lys) that had been subjected to lysine starvation for 30 min (lane −) and from cells not subjected to this nutrient stress (lane +) were annealed with the PpdhA-R primer (see Table S1 in the supplemental material), and then primer extension was performed, as described in Materials and Methods. Lanes G, A, T, and C contained the products of the corresponding dideoxy sequencing reactions performed with the PCR product as the template, as described in Materials and Methods. The nucleotide sequence of the coding strand corresponding to the ladder is shown; the transcription initiation base (+1) is indicated by bold type, and the −35 and −10 regions of the pdhABCD promoter are underlined. The position of the runoff cDNA is indicated by the arrow on the right.