Figure 4.

In vitro and in vivo experimental validation of DasR/PTS^Nag cis/trans relationship. (A) Electrophoretic mobility shift assays. A single HutC/GntR-binding site was present within probes I and II while two were identified in probes III and IV. Probe V, containing the predicted cis-element of CRP of S.coelicolor, was used as control. Probe VI contains the predicted cis-element upstream dasR that presented the best score against the ‘PTScoeli’ weight matrix (see Table 1). A DasR/DNA complex formation was observed for all tested HutC-like cis sites (probes I to IV). (B) In trans effect of DasR on N-acetylglucosamine transport. Mycelia of M145 (pUWL-SK+), the control, and M145 (pFT241 dasR⁺), the DasR overproducing strain, were grown on 50 mM glycerol in the presence and absence of 50 mM N-acetylglucosamine. Data points to determine the initial uptake rates were collected as triplicates and the experiment was reproduced three times. Error bars indicate the derived standard deviations.