Molecular interactions in the control region of the carAB operon encoding Escherichia coli XXXX synthetase.

@entity12116 @entity2036 @entity534 @entity6245 @entity369
The control region of the carAB operon, encoding @entity12116 synthetase, comprises two tandem promoters (P1, upstream and P2, downstream) located 67 base-pairs apart and repressed respectively by @entity6245 and @entity369 . RNA polymerase and pure @entity369 repressor bind to the P2 region in mutually exclusive ways. Repressor protects the two adjacent palindromic @entity534 boxes overlapping P2 against DNase I. Binding of RNA polymerase to P1 is abnormal; the region protected against DNase I is shifted upstream by about 20 @entity2036 with respect to the position expected from the transcription startpoint. This pattern is not due to interference with polymerase binding at P2, since it is observed also in the presence of repressor and on an isolated P1 region. Binding of RNA polymerase is relatively weak and heparin-sensitive suggesting that, in vivo, an ancillary factor is required to promote the formation of an open complex. S1 nuclease mapping experiments show that the simultaneous presence of @entity6245 and @entity369 represses the downstream @entity369 -specific promoter (P2) more efficiently than @entity369 alone. This effect is not due to a direct regulatory interaction between @entity6245 and P2, since it is not observed when P1 is inactivated by insertion mutations or partial deletion. It has been shown that transcription initiated at P1 can proceed even when @entity369 represses P2. We therefore suggest that P2 operator- @entity369 repressor complex is destabilized by RNA polymerase binding at P1 or transcription from P1. We describe a novel technique to select for expression-down mutants in a lac fusion context.
