
Figure 3
Test case: a Notch1 transcription complex containing the RAM region. (a) The first 4.27 Å lowresolution structure of a human Notch1 complex consisting of the Notch ankyrinrepeat domain, the CSL transcription factor and the Mastermindlike 1 (MAML1) coactivator was determined by combining molecular replacement and selenomethionine scanning. Single LeutoMet or ValtoMet mutations (labeled with the MAML1 residue number) were introduced into the MAML1 polypeptide for incorporation of selenomethionine. Anomalous Fourier difference maps were calculated for each of five mutants (the highresolution limit for each data set was between 6 and 7.5 Å) using the anomalous signal from selenomethionine and the phase calculated by molecular replacement. Each map shows a clear peak at the predicted location of the mutated residue, indicated by the matching colors. The gray mesh represents the density for the MAML1 as part of a 2F_{o}  F_{c} density map calculated without any atoms modeled for the MAML1 helix. Adapted from the supplementary information in Nam et al. (2006). (b) An R_{free} heat map of results from the Notch protein DEN optimization using an initial starting temperature of 1000 K. The minimal R_{free} values for each parameter pair (_{DEN} and _{DEN}) over multiple refinement repeats are shown. (c) The corresponding Ramachandran statistics (percentage of disallowed backbone angles). For each parameter pair, the structure with the lowest R_{free} value was usen to calculate the Ramachandran statistics. (d) A histogram showing all calculated R_{free} for a complete portal DEN optimization of the Notch1 complex. The dashed line is the best that could be achieved without DEN (i.e., the lowest R_{free} of the all the `no DEN' repeat refinements); the lowest (or best) R_{free} chosen is the far left tail of the histogram. 