ARM-DOMAIN INTERACTIONS IN ARAC

N-terminal deletions extending beyond the sixth amino acid of the Esch erichia roll regulator of the L-arabinose operon, AraC, were found to generate constitutive regulatory behavior of the promoter p(BAD). Muta genesis of the DNA coding for the first 20 amino acids of the protein and screening for constitutives yielded mutants across the region wher eas screening for mutants tl-lat cannot induce p(BAD), even in the pre sence of arabinose, yielded none. These results indicate that the N-te rminal arm is not essential for transcription activation, but that it plays an important and active role in holding the system in a non-acti vating state. Despite the fact that arabinose binds to the N-terminal domain of AraC, mutations were found in the C-terminal domain that wea ken the binding of arabinose to the protein. The effects of the mutati ons could be suppressed by specific mutation in the N-terminal arm or by deletion of the arm. These results, in conjunction with the crystal structures of the N-terminal domain determined in the presence and ab sence of arabinose, indicate that in the absence of arabinose, the N-t erminal arms of the protein bind to tk le C-terminal DNA binding domai ns to hold them in a state where the protein prefers to loop. When ara binose is added, the arms are pulled off the C-terminal domains, there by releasing them to bind to adjacently located DNA half-sites and act ivate transcription. (C) 1998 Academic Press Limited.