Figure 2

Effect of UV exposure, and presence of CRP-cAMP complex, and additives on integrity of DNA sequence. Panel A: Lane marked “S”: A/G sequence. Lane 1: ³²P-labelled DNA sequence exposed to 15 min of UV exposure (as indicated in Fig. 1, panel A), in the absence of CRP, in buffer (10 mM Tris–HCl, pH 8.0, 50 mM KCl, 2.5 mM MgCl₂, 1 mM EDTA, 55 µg mL⁻¹ bovine serum albumin, 1 mM DTT, 0.05% NP-40, 2 µg mL⁻¹ calf thymus DNA and 50 µM cAMP). Lane 2; as lane 1, except in the presence of 20 nM CRP. Panel B. Representative densitometric scans of lanes 1 and 2 from panel A. Panel C. Lanes marked “S”: A/G sequence. Lane 1: as panel A, but with ³²P-labelled DNA sequence incubated for 15 min in the dark with no CRP; lane 2: as lane 1 but with 20 nM CRP; lane 3: as lane 1 but with UV exposure for 15 min; lane 4: as lane 3, but with 20 nM CRP and UV exposure; lane 5: as lane 4 but in the presence of 100 mM NaN₃; lane 6: as lane 4, but with the omission of DTT from the reaction buffer; lane 7: as lane 4, but in the presence of 100 mM mannitol. Panel D. Sequence (underlined) of the CRP binding site on DNA, with the observed sites of cleavage indicated in red.