FIGURE 7.

In vivo inhibition of cAMP formation in E. coli by serine and aspartate via mutated and reconstituted Tsr and Tar receptor chimeras. A, MacConkey maltose agar plates with E. coli streaked across and a filter strip soaked with 50 μl of either 1 m serine (above) or 1 m aspartate (below) placed on top. Tsr-HAMP₃₆₄₅-Rv3645 AC and Tar-HAMP₃₆₄₅-Rv3645 AC were transformed in BTH101 and DHM1 cells, respectively (see “Experimental Procedures”). The asterisk indicates the faint streak of an E. coli cya strain transformed with a control plasmid (left). At the right, single point mutants expressed individually or concomitantly are probed on a MacConkey agar plate as indicated using DHM-1 cells. The paper strip contained 50 μl of 1 m l-serine. Bottom, control cultures of E. coli wild type and E. coli cya⁻ strains BTH101- and DHM1 are shown. B, β-galactosidase expression is inhibited in cultures by the concomitant presence of either serine (squares) or aspartate (circles) in cells transformed with Tsr-HAMP₃₆₄₅-Rv3645 AC (squares) or Tar-HAMP₃₆₄₅-Rv3645 AC (circles), respectively. Activity of 100% corresponds to β-galactosidase activities without amino acid addition to the culture. Expression was for 90 min. Data were normalized to permit presentation of different β-galactosidase activities in a single graph (n = 3).