FURTHER GENETIC-ANALYSIS OF THE ACTIVATION FUNCTION OF THE TYRR REGULATORY PROTEIN OF ESCHERICHIA-COLI

Previous reports (J, Cui and R, L, Somerville, J, Bacteriol, 175:1777- 1784, 1993; J, Yang, H, Camakaris, and A, J, Pittard, J, Bacteriol. 17 5:6372-6375, 1993) have identified a number of amino acids in the N-te rminal domain of the TyrR protein which are critical for activation of gene expression but which play no role in TyrR-mediated repression. T hese amino acids were clustered in a single region involving positions 2, 3, 5, 7, 9, 10, and 16. Using random and site-directed mutagenesis , we have identified an additional eight key amino acids whose substit ution results in significant or total loss of activator function. All of these are located in the N-terminal domain of TyrR. Alanine scannin g at these eight new positions and at five of the previously identifie d positions for which alanine substitutions had not been obtained has identified three amino acids whose side chains are critical for activa tion, namely, D-9, R-10, and D-103. Glycine at position 37 is also of critical importance. Alanine substitutions at four other positions (C- 7, E-16, D-19, and V-93) caused partial but significant loss of activa tion, indicating that the side chains of these amino acids also play a contributing role in the activation process.