Karen M. Davies, Ed D. Lowe, Catherine Vénien-Bryan and Louise N. Johnson

Supplementary Material

The HupR receiver domain crystal structure in its non-phospho and inhibitory phospho- states

Karen M. Davies, Ed D. Lowe, Catherine Vénien-Bryan and Louise N. Johnson

Figure S1. Sequence alignment of the ATPase-like domain of HupR from Rhodobacter capsulatus with the ATPase domains of other response regulators showing interaction and functional regions. The first four letters indicate the acronym for the activator and the last five letters indicate the organism. Conserved sequences for activators of s54-RNAP have seven regions of conserved sequences designated C1 through C7 (orange boxes). C1 is the Walker A motif, which in Rhodobacter capsulatus HupR and NtrC is not conserved. The lysine and glutamate of the Walker A motif are replaced by arginine and alanine in HupR. C2 in Aquifex aeolicus NtrC1 participates in contacts of the isolated ATPase domain heptamers, as does the region C5.1 C3 carries the GAFTGA motif that is important for coupling energy from ATP hydrolysis to s54 conformational changes. This region is not conserved in HupR, which is expected to uses the s70 factor. C4 is the Walker B motif that contacts nucleotides. The critical aspartate that acts as a catalytic base is replaced by alanine in HupR. Region C6 carried the putative sensor I region and the arginine finger. In NtrC1, either Arg293 or Arg299 could contribute to the catalytic site1. The R finger may also be involved in nucleotide dependent hexamerisation.2 The first arginine is conserved in HupR but not the second, which is replaced by alanine. Region C7 carries the sensor II helix and in NtrC1 Arg357 contacts the b phosphate of ADP. This arginine is conserved in HupR. Red letters indicate conserved residues and blue partially conserved residues.

References

1. Lee, S. Y., DeLaTorre, A., Yan, D., Kustu, S., Nixon, B. T. & Wemmer, D. (2003). Regulation of the Transcriptional Activator NtrC1: Structural Studies of the Regulatory and AAA+ ATPase Domains. Genes Dev 17, 2552-2563.

2. Rappas, M., Schumacher, J., Niwa, H., Buck, M. & Zhang, X. (2006). Structural Basis of the Nucleotide Driven Conformational Changes in the AAA+ Domain of Transcriptional Activator PspF. J Mol Biol 357, 481-492.