Pre-Steady-State Kinetic Analysis of Truncated and Full-Length Saccharomyces cerevisiae DNA Polymerase Eta
Figure 3
Concentration dependence on the pre-steady-state rate constant of nucleotide incorporation catalyzed by truncated yPol. (a) A preincubated solution of truncated yPol (150 nM) and -[32P]-labeled D-7 DNA (30 nM) was mixed with dATPMg2+ (0.4 M, ⬤; 0.8 M, ⚪; 2 M, ■; 4 M, □; 8 M, ▲; 16 M, △; 40 M, ◆; 80 M, ◊) and quenched with EDTA at various time intervals. The solid lines are the best fits to a single-exponential equation which determined the observed rate constant, . (b) The values were plotted as a function of dATP concentration. The data (⬤) were then fit to a hyperbolic equation, yielding a of 6.9 ± 0.4 and a Kd of 17 ± 3 M.
