Fig. 5
The Pgm and PgmL CRDs have no DNA binding activity. a DRaCALA assays using a radiolabeled 80-bp double-strand substrate carrying IES 51A1835 (28 bp) and purified GST or MBP N-terminal protein fusions. Loading onto the membrane was performed in duplicate for all complexes, except for the controls incubated without protein (no prot) or, on the left panel, with the GST tag alone, which were loaded only once. b EMSA assays were performed with a radiolabeled 70-bp double-strand substrate carrying 31 bp from the left end of IES 51A4404 (77 bp) and its left flanking MAC-destined sequences (left), or the same substrate as in panel a for IES 51A1835 (right). c NMR evidence for the lack of an interaction between the Pgm CRD and DNA. Aromatic regions of the 1H-1H NOESY spectra of Pgm(692–768) (black) are not perturbed in the presence of double-stranded DNA (red) from IES 1835 (left) or the left flanking region of IES 51A4404 (right). The sequences of the double-strand DNA substrates used in panels a, b and c are shown in Fig. S3. d Electrostatic potential calculated on one structure of Pgm(701–749). The red and blue colors in surface representations highlight negative and positive charges, respectively