The specific methylation site for Alu I methylase was the cytosine nucleotide in Alu I sequence. The position of the methylated cytosine nucleotide was determined by the chemical cleavage reactions of the Maxam-Gilbert DNA sequencing procedure. As experted, the methylated cytosine nucleotide bands were disappeared on C + T and C lanes on 12% sequencing gels.
Alu I methylase was maximally active at near pH 7.5 in the presence of 50 mM NaCl. The methylase did not require Mg++ for activity, and obeyed Michaelis-Menten Kinetics with respect to both AdoMet and DNA. At 37