Figure 5
Product analysis of by quantitive HPLC. Purified Vh Nag2 (50 ng) was added to a reaction mixture containing 5 mM (GlcNAc)n in 0.2 M mM phosphate buffer, pH 7.0. The mixture was quenched after the indicated reaction times at 30°C by the addition of 0.1 M NaOH and applied to calibrated HPLC. For each substrate (solid line), the calculated concentrations of the products formed (broken line) are shown. Plots show the hydrolysis of A) (GlcNAc)2; B) GlcNAc3; C) GlcNAc4; D) GlcNAc5; E) GlcNAc6; and F) the estimated hydrolytic rate of each substrate as a function of time. Substrates and products are represented by (GlcNAc) (black circle); (GlcNAc)2 (black square); (GlcNAc)3 (black triangle); (GlcNAc)4 (black diamond); (GlcNAc)5 (white square); and (GlcNAc)6 (white triangle).