Reductive methylation of the lysyl residues in the fd gene 5 DNA-binding protein: CD and 13C NMR results on the modified protein.

The epsilon-amino groups of the six lysyl residues of the fd gene 5 DNA-binding protein have been modified by reductive methylation to form N epsilon, N epsilon-dimethyl lysyl derivatives containing 13C-labeled methyl groups. The alpha-amino terminus of the protein was not accessible to methylation. Circular dichroism studies show that the modified protein binds to fd DNA, but with a slightly reduced affinity compared with that of unmodified gene 5 protein. We also find that both the modified and unmodified proteins bind to an oligodeoxynucleotide, d(A)7, but in neither case does binding cause a decrease in the 228 nm CD band of the protein as occurs when the protein binds to long DNA polymers. 13C NMR spectra at 50.1 MHz of [13C]methylated gene 5 protein show five distinct resonances between 43.30 and 42.76 ppm originating from the six N epsilon, N epsilon-dimethyl lysyl residues. We attribute one of the resonances to two solvated lysyl residues and the other four to individual lysyl residues in different microenvironments. All four of these latter resonances are affected by the binding of d(A)7. However, since two of these resonances are similarly affected by the presence of salt in the absence of DNA, only two are uniquely affected by DNA binding.