For all three resistance-conferring mutations in rpoB the value of DG vdW when rifampicin is bound is significantly greater than the same transition for the apo protein and it is this that is mainly driving the positive value of DDG. The difference between the apo-and rifampicin-bound vdW transitions for V170F, S450L and I491F are 4.6, 5.6 and 5.3 kcal/mol, respectively. Since all three of these mutations involve the introduction of a larger sidechain that is oriented towards the bound drug, this is consistent with resistance arising primarily through steric hindrance of the rifampicin binding site. For comparison, despite a similar number of atoms being perturbed, there was no difference in the apo-and drug-bound values of DG vdW for the susceptible mutation rpoB L443F, which is also in the RRDR (Fig. ) and, whilst this also involves the introduction of a larger sidechain, in the crystal structure this is directed away from rifampicin. Differences in DG vdW between the apo-and complexed DNA gyrase also appear mainly responsible for the positive value of DDG for gyrA A90V, however the net effect is reduced. transitions. All results are normalized to the mean of the calculations for the apo leg for each transition for each mutation. Mean values are denoted by a cross and the error bars describe the 95% confidence limits, calculated from the SEM using the appropriate t-statistic. The free energy cost of removing the restraints for rifampicin is not shown since for all mutations it is negligible, indicating that restraints were likely not required to keep the drug in the binding site. An asterisk (*) indicates a gyrB mutation.