FOLDING TOPOLOGY AND DNA-BINDING OF THE N-TERMINAL FRAGMENT OF ADA PROTEIN

Three amino terminal fragments of Escherichia coli Ada protein (39 kDa) with different molecular masses (14 kDa, 16 kDa and 20 kDa) were prepared in large quantities from an E. coli strain harboring plasmids constructed for the overproduction of the truncated proteins. The three fragments can be methylated to an extent similar to that of the intact molecule, The methylated 16 kDa fragment specifically binds to the ada box on a DNA duplex. NMR analyses revealed that the 14 kDa fragment comprises two alpha-helices and a beta-sheet with parallel and anti-parallel mixed strands. A comparison of the N-15-H-1 HMQC spectra of the fragments has led to the conclusion that this tertiary structure within the 14 kDa fragment is retained in the larger 16 kDa and 20 kDa fragments.